Chitin Biosynthesis Research Articles

Evidence of a new intermediate compound of the chitin biogenesis found in the marine-derived Penicilliumroqueforti fungus

Chitin derivatives, chitosan and substituted chito-oligosaccharides from fungi present a wide spectrum of applications and they have been studied in many fields such as medicine, cosmetics, agriculture, aquaculture, and food as dietary supplements [1]. Chitin is a copolymer with N-acetyl-D-glucosamine units linked with β-(1-4)-glycosidic bonds which provide rigidity to the cell wall in chitinous fungi. There is evidence that the chitin synthesis is catalyzed by the chitin synthase (CS; EC 2.4.1.16), an enzyme that transfers β-1,4-linked anhydro-2-acetamido-2-deoxy-D-glucose (GlcNAc) from uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) to the nonreducing end of growing chitin chains [2]. However, chitin is synthesized by the regulation of distinct isoenzymes whose number ranges in some hyphomycetes [3]. Nevertheless, there is relatively little information on the genes responsible for chitin biosynthesis in filamentous fungi, analyses of DNA fragments from taxonomically diverse fungal species have shown that most fungi have three to six chitin synthase genes [4]. Therefore, this diversity of chitin synthases makes difficult to find a unique model of regulation of the chitin pathway. The chemical screening of the biomass of a new marine-derived strain of Penicilliumroqueforti, produced by liquid-state fermentation, led to the identification of several volatile and non-volatile compounds [5]. As a result of this previous study, we have isolated and characterized a new molecule. The chemical structure of the 2-deoxy-2-phosphamino-α-D-glucopyranose isolated was elucidated on the basis of 1D and 2D NMR studies as well as other instrumental techniques. In consequence of this discovery, it has been proposed a biogenetic route aiming to explain its formation as an intermediary component of the chitin biosynthesis.

Role of scalloped in the post-embryonic development of the red flour beetle Tribolium castaneum (Coleoptera: Tenebrionidae)

scalloped (sd) plays a key role in concert with vestigial (vg) in wing development in Drosophila melanogaster Meigen. We previously showed that as well as its conserved role in wing formation in the 28-spotted ladybird beetle Henosepilachna vigintioctopunctata (Fabricius), sd has a novel function in pupal ecdysis. However, the molecular mechanism responsible remains unknown. To determine whether the function of sd in pupal ecdysis was conserved among Coleoptera, we performed expression and functional analyses of sd in the red flour beetle, Tribolium castaneum (Herbst). T. castaneum sd (Tc-sd) mRNA was present continuously from the embryonic stage to the adult. The RNAi-mediated knockdown of Tc-sd in the larval stage resulted in arrest either in the intermolt phase or during larva-to-larva molting. Prepupae injected with Tc-sd dsRNA were arrested as pupae with shorter wings, while its injection in the early pupal stage resulted in arrest as pharate adults. These results suggested that Tc-sd may be involved in growth, molting, and wing formation. The transcript levels of the genes involved in ecdysone biosynthesis, the ecdysone cascade, and chitin biosynthesis were not markedly affected by the knockdown of Tc-sd, which indicated that Tc-sd knockdown did not affect ecdysone biosynthesis or its cascade directly.

Evidence of a New Intermediate Compound of the Chitin Biogenesis Found in a Marine-Derived Fungus

The chemical screening of the biomass of a new marine-derived strain of Penicillium roqueforti, produced by liquid-state fermentation, led to the identification of several volatile and nonvolatile compounds. As a result of previous studies, we have isolated and characterized a new molecule. The chemical structure of 2-deoxy-2-phosphamino-α-D-glucopyranose isolated was elucidated on the basis of 1D and 2D NMR studies together with other instrumental techniques. As a result of this discovery, a biogenetic route has been proposed to explain its formation as an intermediary component of the chitin biosynthesis.

Deciphering the role of the chitin synthase families 1 and 2 in the in vivo and in vitro growth of Aspergillus fumigatus by multiple gene targeting deletion.

Although chitin is an essential component of the fungal cell wall (CW), its biosynthesis and role in virulence is poorly understood. In Aspergillus fumigatus, there are eight chitin synthase (CHS) genes belonging to two families CHSA-C, CHSG in family 1 and CHSF, CHSD, CSMA, CSMB in family 2). To understand the function of these CHS genes, their single and multiple deletions were performed using β-rec/six system to be able to delete all genes within each family (up to a quadruple ΔchsA/C/B/G mutant in family 1 and a quadruple ΔcsmA/csmB/F/D mutant in family 2). Radial growth, conidiation, mycelial/conidial morphology, CW polysaccharide content, Chs-activity, susceptibility to antifungal molecules and pathogenicity in experimental animal aspergillosis were analysed for all the mutants. Among the family 1 CHS, ΔchsA, ΔchsB and ΔchsC mutants showed limited impact on chitin synthesis. In contrast, there was reduced conidiation, altered mycelial morphotype and reduced growth and Chs-activity in the ΔchsG and ΔchsA/C/B/G mutants. In spite of this altered phenotype, these two mutants were as virulent as the parental strain in the experimental aspergillosis models. Among family 2 CHS, phenotypic defects mainly resulted from the CSMA deletion. Despite significant morphological mycelial and conidial growth phenotypes in the quadruple ΔcsmA/csmB/F/D mutant, the chitin content was poorly affected by gene deletions in this family. However, the entire mycelial cell wall structure was disorganized in the family 2 mutants that may be related to the reduced pathogenicity of the quadruple ΔcsmA/csmB/F/D mutant strain compared to the parental strain, in vivo. Deletion of the genes encompassing the two families (ΔcsmA/csmB/F/G) showed that in spite of being originated from an ancient divergence of fungi, these two families work cooperatively to synthesize chitin in A. fumigatus and demonstrate the essentiality of chitin biosynthesis for vegetative growth, resistance to antifungal drugs, and virulence of this filamentous fungus.

PARASITOID VENOM INDUCES METABOLIC CASCADES IN FLY HOSTS.

Parasitoid wasps inject insect hosts with a cocktail of venoms to manipulate the physiology, development, and immunity of the hosts and to promote development of the parasitoid offspring. The jewel wasp Nasonia vitripennis is a model parasitoid with at least 79 venom proteins. We conducted a high-throughput analysis of Nasonia venom effects on temporal changes of 249 metabolites in pupae of the flesh fly host (Sarcophaga bullata), over a five-day time course. Our results show that venom does not simply arrest the metabolism of the fly host. Rather, it targets specific metabolic processes while keeping hosts alive for at least five days post venom injection by the wasp. We found that venom: (a) Activates the sorbitol biosynthetic pathway while maintaining stable glucose levels, (b) Causes a shift in intermediary metabolism by switching to anaerobic metabolism and blocking the tricarboxylic acid cycle, (c) Arrests chitin biosynthesis that likely reflects developmental arrest of adult fly structures, (d) Elevates the majority of free amino acids, and (e) May be increasing phospholipid degradation. Despite sharing some metabolic effects with cold treatment, diapause, and hypoxia, the venom response is distinct from these conditions. Because Nasonia venom dramatically increases sorbitol levels without changing glucose levels, it could be a useful model for studying the regulation of the sorbitol pathway, which is relevant to diabetes research. Our findings generally support the view that parasitoid venoms are a rich source of bioactive molecules with potential biomedical applications.

Rapidly Progressive Lactic Acidosis in Patients with Flufenoxuron Poisoning

Flufenoxuron is an insect growth regulator which inhibits chitin (exoskeleton) biosynthesis in insect's body. Not much is known about its toxicity to human. This report describes three patients who developed worsening lactic acidosis and hypotension after ingestion of flufenoxuron-containing materials. Two of the three patients received continuous renal replacement therapy and survived. (Hong Kong j.emerg.med. 2014;21:181-184)

Insecticide Rotations for the Management of Lepidopteran Pests in Cabbage and Collards

Insecticide rotations for the management of the diamondback moth, Plutella xylostella (L), and other lepidopteran pests of cabbage and collard crops were evaluated in 2010, 2011, and 2012 at Tifton, GA, with the intent of identifying mode of action rotations equal in efficacy to single insecticide mode of action treatments. This was to demonstrate how insecticide rotations, which are purported to aid in the prevention of insecticide resistance, can be an economically-viable insect control option in Brassica crops. The insecticides and mode of actions tested included: chlorantraniliprole and flubendiamide (ryanodine receptor modulators), spinetoram (nicotinic acetylcholine receptor allosteric activator), indoxacarb (voltage-dependent sodium channel blocker), Bacillus thuringiensis subsp. aizawai (microbial disrupter of insect midgut membranes), novaluron (inhibitor of chitin biosynthesis) and zeta-cypermethrin (sodium channel modulator). The results demonstrated that all rotations were equally effective in the control lepidoteran pests in Brassica crops to the use of single insecticide treatments and that at least one rotation treatment provided the highest marketable yield each year.

Regulation of chitin synthesis in pathogenic fungi：an update

Chitin,an essential component of the cell wall,exists widely in pathogenic fungi. Chitin synthase( CHS) is a key enzyme for biosynthesis of chitin,and its activity is strictly regulated; therefore it can be inferred that CHS and its regulation factors are potential targets of anti-fungal therapy. This article reviews the recent research progress on classification,function,and regulation of CHS,and discusses the possibilities of using CHS and its regulation factors as anti-fungal targets.

Sensitivity of Aspergillus nidulans to the Cellulose Synthase Inhibitor Dichlobenil: Insights from Wall-Related Genes’ Expression and Ultrastructural Hyphal Morphologies

The fungal cell wall constitutes an important target for the development of antifungal drugs, because of its central role in morphogenesis, development and determination of fungal-specific molecular features. Fungal walls are characterized by a network of interconnected glycoproteins and polysaccharides, namely α-, β-glucans and chitin. Cell walls promptly and dynamically respond to environmental stimuli by a signaling mechanism, which triggers, among other responses, modulations in wall biosynthetic genes’ expression. Despite the absence of cellulose in the wall of the model filamentous fungus Aspergillus nidulans, we found in this study that fungal growth, spore germination and morphology are affected by the addition of the cellulose synthase inhibitor dichlobenil. Expression analysis of selected genes putatively involved in cell wall biosynthesis, carried out at different time points of drug exposure (i.e. 0, 1, 3, 6 and 24 h), revealed increased expression for the putative mixed linkage β-1,3;1,4 glucan synthase celA together with the β-1,3-glucan synthase fksA and the Rho-related GTPase rhoA. We also compared these data with the response to Congo Red, a known plant/fungal drug affecting both chitin and cellulose biosynthesis. The two drugs exerted different effects at the cell wall level, as shown by gene expression analysis and the ultrastructural features observed through atomic force microscopy and scanning electron microscopy. Although the concentration of dichlobenil required to affect growth of A. nidulans is approximately 10-fold higher than that required to inhibit plant cellulose biosynthesis, our work for the first time demonstrates that a cellulose biosynthesis inhibitor affects fungal growth, changes fungal morphology and expression of genes connected to fungal cell wall biosynthesis.

Trehalose 6-phosphate phosphatase is required for development, virulence and mycotoxin biosynthesis apart from trehalose biosynthesis in Fusarium graminearum

Trehalose 6-phosphate synthase (TPS1) and trehalose 6-phosphate phosphatase (TPS2) are required for trehalose biosynthesis in yeast and filamentous fungi, including Fusarium graminearum. Three null mutants Δtps1, Δtps2 and Δtps1–Δtps2, each carrying either a single deletion of TPS1 or TPS2 or a double deletion of TPS1–TPS2, were generated from a toxigenic F. graminearum strain and were not able to synthesize trehalose. In contrast to its reported function in yeasts and filamentous fungi, TPS1 appeared dispensable for development and virulence. However, deletion of TPS2 abolished sporulation and sexual reproduction; it also altered cell polarity and ultrastructure of the cell wall in association with reduced chitin biosynthesis. The cell polarity alteration was exhibited as reduced apical growth and increased lateral growth and branching with increased hyphal and cell wall widths. Moreover, the TPS2-deficient strain displayed abnormal septum development and nucleus distribution in its conidia and vegetative hyphae. The Δtps2 mutant also had 62% lower mycelial growth on potato dextrose agar and 99% lower virulence on wheat compared with the wild-type. The Δtps1, Δtps2 and Δtps1–Δtps2 mutants synthesized over 3.08-, 7.09- and 2.47-fold less mycotoxins, respectively, on rice culture compared with the wild-type. Comparative transcriptome analysis revealed that the Δtps1, Δtps2 and Δtps1–Δtps2 mutants had 486, 1885 and 146 genotype-specific genes, respectively, with significantly changed expression profiles compared with the wild-type. Further dissection of this pathway will provide new insights into regulation of fungal development, virulence and trichothecene biosynthesis.

Transcriptome Analysis of Encystation in Entamoeba invadens

Encystation is an essential differentiation process for the completion of the life cycle of a group of intestinal protozoa including Entamoeba histolytica, the causative agent of intestinal and extraintestinal amebiasis. However, regulation of gene expression during encystation is poorly understood. To comprehensively understand the process at the molecular level, the transcriptomic profiles of E . invadens , which is a related reptilian species that causes an invasive disease similar to that of E. histolytica, was investigated during encystation. Using a custom-generated Affymetrix platform microarray, we performed time course (0.5, 2, 8, 24, 48, and 120 h) gene expression analysis of encysting E . invadens . ANOVA analysis revealed that a total of 1,528 genes showed ≥3 fold up-regulation at one or more time points, relative to the trophozoite stage. Of these modulated genes, 8% (116 genes) were up-regulated at the early time points (0.5, 2 and 8h), while 63% (962 genes) were up-regulated at the later time points (24, 48, and 120 h). Twenty nine percent (450 genes) are either up-regulated at 2 to 5 time points or constitutively up-regulated in both early and late stages. Among the up-regulated genes are the genes encoding transporters, cytoskeletal proteins, proteins involved in vesicular trafficking (small GTPases), Myb transcription factors, cysteine proteases, components of the proteasome, and enzymes for chitin biosynthesis. This study represents the first kinetic analysis of gene expression during differentiation from the invasive trophozoite to the dormant, infective cyst stage in Entamoeba . Functional analysis on individual genes and their encoded products that are modulated during encystation may lead to the discovery of targets for the development of new chemotherapeutics that interfere with stage conversion of the parasite.

Molecular and Functional Analysis of UDP-N-Acetylglucosamine Pyrophosphorylases from the Migratory Locust, Locusta migratoria

UDP-N-acetylglucosamine pyrophosphorylases (UAP) function in the formation of extracellular matrix by producing N-acetylglucosamine (GlcNAc) residues needed for chitin biosynthesis and protein glycosylation. Herein, we report two UAP cDNA’s derived from two different genes (LmUAP1 and LmUAP2) in the migratory locust Locusta migratoria. Both the cDNA and their deduced amino acid sequences showed about 70% identities between the two genes. Phylogenetic analysis suggests that LmUAP1 and LmUAP2 derive from a relatively recent gene duplication event. Both LmUAP1 and LmUAP2 were widely expressed in all the major tissues besides chitin-containing tissues. However, the two genes exhibited different developmental expression patterns. High expression of LmUAP1 was detected during early embryogenesis, then decreased greatly, and slowly increased before egg hatch. During nymphal development, the highest expression of LmUAP1 appeared just after molting but declined in each inter-molting period and then increased before molting to the next stage, whereas LmUAP2 was more consistently expressed throughout all these stages. When the early second- and fifth-instar nymphs (1-day-old) were injected with LmUAP1 double-stranded RNA (dsRNA), 100% mortality was observed 2 days after the injection. When the middle second- and fifth-instar nymphs (3- to 4-day-old) were injected with LmUAP1 dsRNA, 100% mortality was observed during their next molting process. In contrast, when the insects at the same stages were injected with LmUAP2 dsRNA, these insects were able to develop normally and molt to the next stage successfully. It is presumed that the lethality caused by RNAi of LmUAP1 is due to reduced chitin biosynthesis of the integument and midgut, whereas LmUAP2 is not essential for locust development at least in nymph stage. This study is expected to help better understand different functions of UAP1 and UAP2 in the locust and other insect species.

Identification and first insights into the structure and biosynthesis of chitin from the freshwater sponge Spongilla lacustris

This work demonstrates that chitin is an important structural component within the skeletal fibers of the freshwater sponge Spongilla lacustris. Using a variety of analytical techniques ((13)C solid state NMR, FT-IR, Raman, NEXAFS, ESI-MS, Morgan-Elson assay and Calcofluor White Staining); we show that this sponge chitin is much closer to α-chitin, known to be present in other animals, than to β-chitin. Genetic analysis confirmed the presence of chitin synthases, which are described for the first time in a sponge. The presence of chitin in both marine (demosponges and hexactinellids) and freshwater sponges indicates that this important structural biopolymer was already present in their common ancestor.

Conserved microRNAs miR-8-5p and miR-2a-3p modulate chitin biosynthesis in response to 20-hydroxyecdysone signaling in the brown planthopper, Nilaparvata lugens

Molting is an important developmental process in insects, usually along with synthesis and degradation of chitin. 20-hydroxyecdysone (20E), an insect hormone, has been reported to contribute to many processes including molting. However, little is known about the link between the chitin biosynthesis pathway and 20E signaling. Here, we report that conserved miR-8-5p (miR-8-5p) and miR-2a-3p and their new target genes are critical for ecdysone-induced chitin biosynthesis in a hemipteran insect Nilaparvata lugens. We found that membrane-bound trehalase (Tre-2) and phosphoacetylglucosamine mutase (PAGM) in the chitin biosynthesis pathway were targets of miR-8-5p and miR-2a-3p, respectively, through bioinformatic analysis and experimental verification. The levels of miR-8-5p and miR-2a-3p were reduced, whereas the levels of Tre-2 and PAGM were up-regulated in response to 20E. In addition, miR-8-5p and miR-2a-3p were transcriptionally repressed by an early-response gene, the Broad-Complex (BR-C), in the 20E signaling pathway. Moreover, the overexpression of miR-8-5p and miR-2a-3p led to a significant reduction in the survival rate along with a molting obstacles defect phenotype caused by miR-2a-3p mimics feeding, and the chitin content of N. lugens was simultaneously reduced. Thus, miR-8-5p and miR-2a-3p act as molecular link that tune the chitin biosynthesis pathway in response to 20E signaling.

Synergy of the antibiotic colistin with echinocandin antifungals in Candida species

Candida albicans is the most prevalent fungal pathogen of humans, causing a wide range of infections from harmless superficial to severe systemic infections. Improvement of the antifungal arsenal is needed since existing antifungals can be associated with limited efficacy, toxicity and antifungal resistance. Here we aimed to identify compounds that act synergistically with echinocandin antifungals and that could contribute to a faster reduction of the fungal burden. A total of 38 758 compounds were tested for their ability to act synergistically with aminocandin, a β-1,3-glucan synthase inhibitor of the echinocandin family of antifungals. The synergy between echinocandins and an identified hit was studied with chemogenomic screens and testing of individual Saccharomyces cerevisiae and C. albicans mutant strains. We found that colistin, an antibiotic that targets membranes in Gram-negative bacteria, is synergistic with drugs of the echinocandin family against all Candida species tested. The combination of colistin and aminocandin led to faster and increased permeabilization of C. albicans cells than either colistin or aminocandin alone. Echinocandin susceptibility was a prerequisite to be able to observe the synergy. A large-scale screen for genes involved in natural resistance of yeast cells to low doses of the drugs, alone or in combination, identified efficient sphingolipid and chitin biosynthesis as necessary to protect S. cerevisiae and C. albicans cells against the antifungal combination. These results suggest that echinocandin-mediated weakening of the cell wall facilitates colistin targeting of fungal membranes, which in turn reinforces the antifungal activity of echinocandins.

Silencing ofEntamoeba histolyticaGlucosamine 6-Phosphate Isomerase by RNA Interference Inhibits the Formation of Cyst-Like Structures

Encystment is an essential process in the biological cycle of the human parasite Entamoeba histolytica. In the present study, we evaluated the participation of E. histolytica Gln6Pi in the formation of amoeba cyst-like structures by RNA interference assay. Amoeba trophozoites transfected with two Gln6Pi siRNAs reduced the expression of the enzyme in 85%, which was confirmed by western blot using an anti-Gln6Pi antibody. The E. histolytica Gln6Pi knockdown with the mix of both siRNAs resulted in the loss of its capacity to form cyst-like structures (CLSs) and develop a chitin wall under hydrogen peroxide treatment, as evidenced by absence of both resistance to detergent treatment and calcofluor staining. Thus, only 5% of treated trophozoites were converted to CLS, from which only 15% were calcofluor stained. These results represent an advance in the understanding of chitin biosynthesis in E. histolytica and provide insight into the encystment process in this parasite, which could allow for the developing of new control strategies for this parasite.

The chitin biosynthesis pathway in Entamoeba and the role of glucosamine-6-P isomerase by RNA interference

Entamoeba histolytica, the causative agent of amoebiasis, infects through its cyst form. A thick chitin wall protects the cyst from the harsh environment outside of the body. It is known that chitin is synthesized only during encystation, but the chitin synthesis pathway (CSP) of Entamoeba is not well characterized. In this report, we have identified the genes involved in chitin biosynthesis from the Entamoeba genome database and verified their expression profile at the transcriptional level in encysting Entamoeba invadens. Semi-quantitative RT-PCR (sqRT-PCR) analysis showed that all the chitin pathway genes are entirely absent or transcribed at low levels in trophozoites. The mRNA expression of most of the CSP genes reached their maximum level between 9 and 12h after the in vitro initiation of encystation. Double-stranded RNA-mediated silencing of glucosamine-6-P isomerase (Gln6Pi) reduced chitin synthesis to 62–64%, which indicates that Gln6Pi might be a key enzyme for regulating chitin synthesis in Entamoeba. The study of different enzymes involved in glycogen metabolism revealed that stored glycogen is converted to glucose during encystation. It is clear from the sqRT-PCR analysis that the rate of glycolysis decreases as encystation proceeds. Encystation up-regulates the expression of glycogen phosphorylase, which is responsible for glycogen degradation. The significant decrease in chitin synthesis in encysting cells treated with a specific inhibitor of glycogen phosphorylase indicates that the glucose obtained from the degradation of stored glycogen in trophozoites might be one of the major sources of glucose for chitin synthesis.

Dramatic Increase in Glycerol Biosynthesis upon Oxidative Stress in the Anaerobic Protozoan Parasite Entamoeba histolytica

Entamoeba histolytica, a microaerophilic enteric protozoan parasite, causes amebic colitis and extra intestinal abscesses in millions of inhabitants of endemic areas. Trophozoites of E. histolytica are exposed to a variety of reactive oxygen and nitrogen species during infection. Since E. histolytica lacks key components of canonical eukaryotic anti-oxidative defense systems, such as catalase and glutathione system, alternative not-yet-identified anti-oxidative defense strategies have been postulated to be operating in E. histolytica. In the present study, we investigated global metabolic responses in E. histolytica in response to H2O2- and paraquat-mediated oxidative stress by measuring charged metabolites on capillary electrophoresis and time-of-flight mass spectrometry. We found that oxidative stress caused drastic modulation of metabolites involved in glycolysis, chitin biosynthesis, and nucleotide and amino acid metabolism. Oxidative stress resulted in the inhibition of glycolysis as a result of inactivation of several key enzymes, leading to the redirection of metabolic flux towards glycerol production, chitin biosynthesis, and the non-oxidative branch of the pentose phosphate pathway. As a result of the repression of glycolysis as evidenced by the accumulation of glycolytic intermediates upstream of pyruvate, and reduced ethanol production, the levels of nucleoside triphosphates were decreased. We also showed for the first time the presence of functional glycerol biosynthetic pathway in E. histolytica as demonstrated by the increased production of glycerol 3-phosphate and glycerol upon oxidative stress. We proposed the significance of the glycerol biosynthetic pathway as a metabolic anti-oxidative defense system in E. histolytica.

Characterization of a midgut-specific chitin synthase gene (LmCHS2) responsible for biosynthesis of chitin of peritrophic matrix in Locusta migratoria

Chitin, an essential component of peritrophic matrix (PM), is produced by a series of biochemical reactions. Chitin synthase plays a crucial role in chitin polymerization in chitin biosynthetic pathway. In this study, we identified and characterized a full-length cDNA of chitin synthase 2 gene (LmCHS2) from Locusta migratoria. The cDNA contains an open reading frame of 4569 nucleotides that encode 1523 amino acid residues, and 76- and 373-nucleotides for 5′- and 3′-noncoding regions, respectively. Analysis of LmCHS2 transcript in different tissues of the locust by using real-time quantitative PCR indicated that LmCHS2 was exclusively expressed in midgut and gastric caeca (a part of the midgut). The highest expression was found in the anterior midgut with a decline of the transcript level from the anterior to posterior regions. During growth and development of locusts, there was only a slight expression in eggs, but the expression gradually increased from nymphs to adults. In situ hybridization further revealed that LmCHS2 transcript mainly presented in the apical regions of brush border forming columnar cells of gastric caeca. LmCHS2 dsRNA was injected to fifth-instar nymphs to further explore biological functions of LmCHS2. Significantly down-regulated transcript of LmCHS2 resulted in a cessation of feeding and a high mortality of the insect. However, no visible abnormal morphological change of locusts was observed until insects molted to adults. After dissection, we found that the average length of midguts from the LmCHS2 dsRNA-injected locusts was shorter than that of the control insects that were injected with dsGFP. Furthermore, microsection of midguts showed that the PM of the LmCHS2 dsRNA-injected nymphs was amorphous and thin as compared with the controls. Our results demonstrate that LmCHS2 is responsible for the biosynthesis of chitin associated with PM and plays an essential role in locust growth and development.

Síntese de inibidores potenciais de hexosaminidases derivados de N-acetilglicosamina e D-glicose

We report herein the synthesis and characterization of N-acetylglucosamine and D-glucose derivates modified at C-1 for evaluation of antifungal activity as potential hexosaminidases inhibitors. There are various related antifungal commercial drugs, but all of them have collateral damages like: irreversible kidney disease, hormonal disorders and antibiotic resistance . In this current scenario the study of new antifungal drugs that can act by a new mechanism is important. Several natural and synthetic hexosaminidases inhibitory are reported in the literature. These enzymes are critical for the biosynthesis and remodeling of chitin, important for fungi structure. Hexosaminidases inhibitors are generally monosaccharides derivates or analogs of N-acetylglucosamine.