Bacteria inhabiting spider webs enhance host silk extensibility.

Spider silk is a promising material with great potential in biomedical applications due to its incredible mechanical properties and resistance to degradation of commercially available bacterial strains. However, little is known about the bacterial communities that may inhabit spider webs and how these microorganisms interact with spider silk. In this study, we exposed two exopolysaccharide-secreting bacteria, isolated from webs of an orb spider, to major ampullate (MA) silk from host spiders. The naturally occurring lipid and glycoprotein surface layers of MA silk were experimentally removed to further probe the interaction between bacteria and silk. Extensibility of major ampullate silk produced by Triconephila clavata that was exposed to either Microbacterium sp. or Novosphigobium sp. was significantly higher than that of silk that was not exposed to bacteria (differed by 58.7%). This strain-enhancing effect was not observed when the lipid and glycoprotein surface layers of MA silks were removed. The presence of exopolysaccharides was detected through NMR from MA silks exposed to these two bacteria but not from those without exposure. Here we report for the first time that exopolysaccharide-secreting bacteria inhabiting spider webs can enhance extensibility of host MA silks and silk surface layers play a vital role in mediating such effects.

Comprehensive analysis of silk proteins and gland compartments in Limnephilus lunatus, a case-making trichopteran.

Caddisfly larvae produce silk containing heavy and light fibroins, similar to the silk of Lepidoptera, for the construction of underwater structures. We analyzed the silk of Limnephilus lunatus belonging to the case-forming suborder Integripalpia. We analyzed the transcriptome, mapped the transcripts to a reference genome and identified over 80 proteins using proteomic methods, and checked the specificity of their expression. For comparison, we also analyzed the transcriptome and silk proteome of Limnephilus flavicornis. Our results show that fibroins and adhesives are produced together in the middle and posterior parts of the silk glands, while the anterior part produces enzymes and an unknown protein AT24. The number of silk proteins of L. lunatus far exceeds that of the web-spinning Plectrocnemia conspersa, a previously described species from the suborder Annulipalpia. Our results support the idea of increasing the structural complexity of silk in rigid case builders compared to trap web builders.

Sustainable Silk-Based Particulate Systems for the Controlled Release of Pharmaceuticals and Bioactive Agents in Wound Healing and Skin Regeneration.

The increasing demand for innovative approaches in wound healing and skin regeneration has prompted extensive research into advanced biomaterials. This review focuses on showcasing the unique properties of sustainable silk-based particulate systems in promoting the controlled release of pharmaceuticals and bioactive agents in the context of wound healing and skin regeneration. Silk fibroin and sericin are derived from well-established silkworm production and constitute a unique biocompatible and biodegradable protein platform for the development of drug delivery systems. The controlled release of therapeutic compounds from silk-based particulate systems not only ensures optimal bioavailability but also addresses the challenges associated with conventional delivery methods. The multifaceted benefits of silk proteins, including their inherent biocompatibility, versatility, and sustainability, are explored in this review. Furthermore, the intricate mechanisms by which controlled drug release takes place from silk-based carriers are discussed.

A silk-based hydrogel containing dexamethasone and lipoic acid microcrystals for local delivery to the inner ear.

Local drug delivery has been exploited recently to treat hearing loss, as this method can both bypass the blood-labyrinth barrier and provide sustained drug release. Combined drug microcrystals (MCs) offer additional advantages for sensorineural hearing loss treatment via intratympanic (IT) injection due to their shape effect and combination strategy. In this study, to endow viscous effects of hydrogels, nonspherical dexamethasone (DEX) and lipoic acid (LA) MCs were incorporated into silk fibroin (SF) hydrogels, which were subsequently administered to the tympanic cavity to investigate their pharmaceutical properties. First, we prepared DEX and LA MCs by a traditional precipitation technique followed by SF hydrogel incorporation (SF+DEX+LA). After characterization of the physicochemical features, including morphology, rheology, and dissolution, both a suspension of combined DEX and LA MCs (DEX+LA) and SF+DEX+LA were administered to guinea pigs by IT injection, after which the pharmacokinetics, biodegradation and biocompatibility were evaluated. To our surprise, compared to the DEX+LA group, the pharmacokinetics of the SF+DEX+LA hydrogel group did not improve significantly, which may be ascribed to their nonspherical shape and deposition effects of the drugs MCs. The cochlear tissue in each group displayed good morphology, with no obvious inflammatory reactions. This combined MC suspension has the clear advantages of no vehicle, easy scale-up preparation, and good biocompatibility and outcomes, which paves the way for practical treatment of hearing loss via local drug delivery.

Harvesting Silk Fibers from Plodia interpunctella: Role of Environmental Rearing Conditions in Fiber Production and Properties.

Silk fibers are produced by a wide variety of insects. The silkworm Bombyx mori (Bombyx) was domesticated because the physical properties of its silk fibers were amenable to the production of fine textiles. Subsequently, engineers have regenerated silk fibroin to form biomaterials. The monocular focus on Bombyx silk has underutilized the expanse of diverse silk proteins produced by more than 100,000 other arthropods. This vast array of silk fibers could be utilized for biomedical engineering challenges if sufficient rearing and purification processes are developed. Herein, we show that the moth, Plodia interpunctella (Plodia), represents an alternative silk source that is easily reared in highly regulated culture environments allowing for greater consistency in the silk produced. We controlled the temperature, resource availability (larvae/gram diet), and population density (larvae/mL) with the goal of increasing silk fiber production and improving homogeneity in Plodia silk proteins. We determined that higher temperatures accelerated insect growth and reduced life cycle length. Furthermore, we established initial protocols for the production of Plodia silk with optimal silk production occurring at 24 °C, with a resource availability of 10 larvae/gram and a population density of 0.72 larvae/mL. Population density was shown to be the most prominent driving force of Plodia silk mat formation among the three parameters assessed. Future work will need to link gene expression, protein production and purification, and resulting mechanical properties as a function of environmental cues to further transition Plodia silk into regenerated silk fibroin biomaterials.

CRISPR/Cas9-Mediated Editing of BmEcKL1 Gene Sequence Affected Silk Gland Development of Silkworms (Bombyx mori).

The silkworm (Bombyx mori) has served humankind through silk protein production. However, traditional sericulture and the silk industry have encountered considerable bottlenecks and must rely on major technological breakthroughs to keep up with the current rapid developments. The adoption of gene editing technology has nevertheless brought new hope to traditional sericulture and the silk industry. The long period and low efficiency of traditional genetic breeding methods to obtain high silk-yielding silkworm strains have hindered the development of the sericulture industry; the use of gene editing technology to specifically control the expression of genes related to silk gland development or silk protein synthesis is beneficial for obtaining silkworm strains with excellent traits. In this study, BmEcKL1 was specifically knocked out in the middle (MSGs) and posterior (PSGs) silk glands using CRISPR/Cas9 technology, and ΔBmEcKL1-MSG and ΔBmEcKL1-PSG strains with improved MSGs and PSGs and increased silk production were obtained. This work identifies and proves that BmEcKL1 directly or indirectly participates in silk gland development and silk protein synthesis, providing new perspectives for investigating silk gland development and silk protein synthesis mechanisms in silkworms, which is of great significance for selecting and breeding high silk-yielding silkworm varieties.

The ratio of fibroin to sericin in the middle silk gland of Bombyx mori and its correlation with the extensional behavior of the silk dope.

Understanding how native silk spinning occurs is crucial for designing artificial spinning systems. One often overlooked factor in Bombyx mori is the secretion of sericin proteins. Herein, we investigate the variation in amino acid content at different locations in the middle silk gland (MSG) of B. mori. This variation corresponds to an increase in sericin content when moving towards the anterior region of the MSG, while the posterior region predominantly contains fibroin. We estimate the mass ratio of sericin to fibroin to be ~25/75 wt% in the anterior MSG, depending on the fitting method. Then, we demonstrate that the improvement in the extensional behavior of the silk dope in the MSG correlates with the increase in sericin content. The addition of sericin may decrease the viscosity of the silk dope, a factor associated with an increase in the spinnability of silk. We further discuss whether this effect could also result from other known physicochemical changes within the MSG.

Nonsteaming method improves the nutritional value and utilization efficiency of silkworm artificial diets.

Artificial diets for silkworms overcome the seasonal limitations of traditional rearing methods with fresh mulberry leaves. However, the current wet artificial diets, steamed at high temperatures, are not favored by silkworms, and they are cumbersome and challenging to preserve. These conditions adversely affected the development of artificial diet-based sericulture production. In this study, we disinfected dry powder diets with radiation and added distilled water without steaming before use. Then, the nutritional value of finished diets and their impact on silkworm development was assessed. Compared with steamed diets, nonsteamed diets were more attractive to silkworms. Chemical assays showed significantly more essential nutrients for silkworms, including l-ascorbic acid, vitamin B1, vitamin B2, and urease in nonsteamed diets than in steamed diets. Feeding fifth-instar silkworm larvae with nonsteamed diets significantly improved the ammonia utilization efficiency of the diet and increased the cocoon shell rate and diet/silk protein conversion efficiency by 5.9% and 13.3%, respectively. When fed with nonsteamed diets, the abundance of aerobic microorganisms in silkworm intestines increased and the abundance of pathogenic bacteria decreased. Furthermore, the vitality of the silkworm, measured by the dead worm cocoon rate, significantly improved by 16.90%. In summary, preparing sterile wet diets without high-temperature steaming effectively improved the nutritional value of the diet and enhanced silkworm growth.

An Advanced Bioconjugation Strategy for Covalent Tethering of TGFß3 with Silk Fibroin Matrices and its Implications in the Chondrogenesis Profile of Human BMSCs and Human Chondrocytes: A Paradigm Shift in Cartilage Tissue Engineering.

The transforming growth factor-ß class of cytokines plays a significant role in articular cartilage formation from mesenchymal condensation to chondrogenic differentiation. However, their exogenous addition to the chondrogenic media makes the protocol expensive. It reduces the bioavailability of the cytokine to the cells owing to their burst release. The present study demonstrates an advanced bioconjugation strategy to conjugate transforming growth factor-ß3 (TGFß3) with silk fibroin matrix covalently via a cyanuric chloride coupling reaction. The tethering and change in secondary conformation are confirmed using various spectroscopic analyses. To assess the functionality of the chemically modified silk matrix, human bone marrow-derived mesenchymal stem cells (hBMSCs) and chondrocytes are cultured for 28 days in a chondrogenic differentiation medium. Gene expression and histological analysis reveal enhanced expression of chondrogenic markers with intense Safranin-O and Alcian Blue staining in TGFß3 conjugated silk matrices than where TGFß3 is exogenously added to the media for both hBMSCs and chondrocytes. Therefore, this study successfully recapitulates the native niche of TGFß3 and the role of the silk as a growth factor stabilizer. When cultured over TGFß3 conjugated silk matrices, hBMSCs display increased proteoglycan secretion and maximum chondrogenic trait with attenuation of chondrocyte hypertrophy over human chondrocytes.

Identification and function of the Pax gene Bmgsb in the silk gland of Bombyx mori.

Paired box (Pax) genes are highly conserved throughout evolution, and the Pax protein is an important transcription factor of embryonic development. The Pax gene Bmgsb is expressed in the silk glands of silkworm, but its biological functions remain unclear. This study aimed to investigate the expression pattern of Bmgsb in the silk gland and explore its functions using RNA interference (RNAi). Here, we identified eight Pax genes in Bombyx mori. Phylogenetic analysis showed that the B. mori Pax genes were highly homologous to the Pax genes in other insects and highly evolutionarily conserved. The tissue expression profile showed that Bmgsb was expressed in the anterior silk gland and anterior part of the middle silk gland (AMSG). RNAi of Bmgsb resulted in defective development of the AMSG, and the larvae were mostly unable to cocoon in the wandering stage. RNA-seq analysis showed that the fibroin genes fib-l, fib-h and p25, cellular heat shock response-related genes and phenol oxidase genes were considerably upregulated upon Bmgsb knockdown. Furthermore, quantitative reverse transcription-PCR results showed that the fibroin genes and ubiquitin proteolytic enzyme-related genes were significantly upregulated in the AMSG after Bmgsb knockdown. This study provides a foundation for future research on the biological functions of B. mori Pax genes. In addition, it demonstrates the important roles of Bmgsb in the transcriptional regulation of fibroin genes and silk gland development.

Production and secretion of recombinant spider silk in Bacillus megaterium.

BACKGROUND: Silk proteins have emerged as versatile biomaterials with unique chemical and physical properties, making them appealing for various applications. Among them, spider silk, known for its exceptional mechanical strength, has attracted considerable attention. Recombinant production of spider silk represents the most promising route towards its scaled production; however, challenges persist within the upstream optimization of host organisms, including toxicity and low yields. The high cost of downstream cell lysis and protein purification is an additional barrier preventing the widespread production and use of spider silk proteins. Gram-positive bacteria represent an attractive, but underexplored, microbial chassis that may enable a reduction in the cost and difficulty of recombinant silk production through attributes that include, superior secretory capabilities, frequent GRAS status, and previously established use in industry. RESULTS: In this study, we explore the potential of gram-positive hosts by engineering the first production and secretion of recombinant spider silk in the Bacillus genus. Using an industrially relevant B. megaterium host, it was found that the Sec secretion pathway enables secretory production of silk, however, the choice of signal sequence plays a vital role in successful secretion. Attempts at increasing secreted titers revealed that multiple translation initiation sites in tandem do not significantly impact silk production levels, contrary to previous findings for other gram-positive hosts and recombinant proteins. Notwithstanding, targeted amino acid supplementation in minimal media was found to increase production by 135% relative to both rich media and unaltered minimal media, yielding secretory titers of approximately 100 mg/L in flask cultures. CONCLUSION: It is hypothesized that the supplementation strategy addressed metabolic bottlenecks, specifically depletion of ATP and NADPH within the central metabolism, that were previously observed for an E. coli host producing the same recombinant silk construct. Furthermore, this study supports the hypothesis that secretion mitigates the toxicity of the produced silk protein on the host organism and enhances host performance in glucose-based minimal media. While promising, future research is warranted to understand metabolic changes more precisely in the Bacillus host system in response to silk production, optimize signal sequences and promoter strengths, investigate the mechanisms behind the effect of tandem translation initiation sites, and evaluate the performance of this system within a bioreactor.

Exploring the apoptotic effects of sericin on HCT116 cells through comprehensive nanostring transcriptomics and proteomics analysis.

Sericin, a silk protein from Bombyx mori (silkworms), has many applications, including cosmetics, anti-inflammation, and anti-cancer. Sericin complexes with nanoparticles have shown promise for breast cancer cell lines. Apoptosis, a programmed cell death mechanism, stops cancer cell growth. This study found that Sericin urea extract significantly affected HCT116 cell viability (IC50 = 42.00 ± 0.002 µg/mL) and caused apoptosis in over 80% of treated cells. S-FTIR analysis showed significant changes in Sericin-treated cells' macromolecule composition, particularly in the lipid and nucleic acid areas, indicating major cellular modifications. A transcriptomics study found upregulation of the apoptotic signaling genes FASLG, TNFSF10, CASP3, CASP7, CASP8, and CASP10. Early apoptotic proteins also showed that BAD, AKT, CASP9, p53, and CASP8 were significantly upregulated. A proteomics study illuminated Sericin-treated cells' altered protein patterns. Our results show that Sericin activated the extrinsic apoptosis pathway via the caspase cascade (CASP8/10 and CASP3/7) and the death receptor pathway, involving TNFSF10 or FASLG, in HCT116 cells. Upregulation of p53 increases CASP8, which activates CASP3 and causes HCT116 cell death. This multi-omics study illuminates the molecular mechanisms of Sericin-induced apoptosis, sheds light on its potential cancer treatment applications, and helps us understand the complex relationship between silk-derived proteins and cellular processes.

Effect of eIF6 on the development of silk glands and silk protein synthesis of the silkworm, Bombyx mori.

The silkworm is a lepidopteran domesticated from the wild silkworm, mostly valued for its efficient synthesis of silk protein. This species' ability to spin silk has supported the 5500-year-old silk industry and the globally known "Silk Road", making the transformation of mulberry leaves into silk of great concern. Therefore, research on the silk-related genes of silkworms and their regulatory mechanisms has attracted increasing attention. Previous studies have revealed that domestic silk gland cells are endoreduplication cells, and their high-copy genome and special chromatin conformation provide conditions for the high expression of silk proteins. In this study, we systematically investigate the expression pattern of eukaryotic initiation factors (eIFs) and identified the eIF6 as a eukaryotic translation initiation factor involved in the synthesis of silk proteins. We generated an eIF6 gene deletion mutant strain of silkworm using the CRISPR/Cas9 system and investigated the function of eIF6 in silk gland development and silk protein synthesis. The results showed that deletion of eIF6 inhibited the individual development of silkworm larvae, inhibited the development of silk glands, and significantly reduced the cocoon layer ratio. Therefore, we elucidated the function of eIF6 in the development of silk glands and the synthesis of silk proteins, which is important for further elucidation of the developmental process of silk glands and the mechanism underlying the ultra-high expression of silk proteins.

"Needle" hidden in silk floss: Inactivation effect and mechanism of melamine sponge loaded bismuth oxide composite copper-metal organic framework (MS/Bi2O3@Cu-MOF) as floating photocatalyst on Microcystis aeruginosa.

Photocatalytic technology showed significant potential for addressing the issue of cyanobacterial blooms resulting from eutrophication in bodies of water. However, the traditional powder materials were easy to agglomerate and settle, which led to the decrease of photocatalytic activity. The emergence of floating photocatalyst was important for the practical application of controlling harmful algal blooms. This study was based on the efficient powder photocatalyst bismuth oxide composite copper-metal organic framework (Bi2O3 @Cu-MOF), which was successfully loaded onto melamine sponge (MS) by sodium alginate immobilization to prepare a floating photocatalyst MS/Bi2O3 @Cu-MOF for the inactivation of Microcystis aeruginosa (M. aeruginosa) under visible light. When the capacity was 0.4 g (CA0.4), MS/Bi2O3 @Cu-MOF showed good photocatalytic activity, and the inactivation rate of M. aeruginosa reached 74.462% after 120 h. MS/Bi2O3 @Cu-MOF-CA0.4 showed a large specific surface area of 30.490 m2/g and an average pore size of 22.862 nm, belonging to mesoporous materials. After 120 h of treatment, the content of soluble protein in the MS/Bi2O3 @Cu-MOF-CA0.4 treatment group decreased to 0.365 mg/L, the content of chlorophyll a (chla) was 0.023 mg/L, the content of malondialdehyde (MDA) increased to 3.168 nmol/mgprot, and the contents of various antioxidant enzymes experienced drastic changes, first increasing and then decreasing. The photocatalytic process generated·OH and·O2-, which played key role in inactivating the algae cells. Additionally, the release of Cu2+ and adsorption of the material also contributed to the process.

Low concentration chlorantraniliprole-promoted Ca2+ release drives a shift from autophagy to apoptosis in the silk gland of Bombyx mori.

The novel pesticide chlorantraniliprole (CAP) is widely used for pest control in agriculture, and the safety for non-target organisms of trace residues in the environment has received widespread attention. In the present study, exposure to low concentrations of CAP resulted in abnormal silk gland development in the B. mori, and induced the release of intracellular Ca2+ in addition to the triggering of Ca2+-dependent gene transcription. Moreover, the CAP treatment group exhibited down-regulation of oxidative phosphorylation and antioxidant enzyme-related genes in the silk gland, resulting in peroxide accumulation. Furthermore, transcript levels of autophagy-related genes were significantly up-regulated and protein levels of LC3-I and LC3-II were up-regulated, indicating an increase in autophagy. The protein levels of ATG5 and NtATG5 were also significantly up-regulated. While the protein levels of caspase3 and active caspase3 were significantly up-regulated consistent with the transcript levels of key genes in the apoptotic signaling pathway, ultimately affecting silk protein synthesis. Overall, these findings indicate that low concentration CAP induced abnormal development in the silk gland of B. mori by causing intracellular Ca2+ overload, which inhibits oxidative phosphorylation pathway and the removal of reactive oxygen species, leading to a driving a shift from autophagy to apoptosis. The findings herein provided a basis for evaluating the safety of CAP environmental residues on non-target organisms.

The promoting effects of pyriproxyfen on autophagy and apoptosis in silk glands of non-target insect silkworm, Bombyx mori.

Pyriproxyfen is a juvenile hormone analogue. The physiological effects of its low-concentration drift during the process of controlling agricultural and forestry pests on non-target organisms in the ecological environment are unpredictable, especially the effects on organs that play a key role in biological function are worthy of attention. The silk gland is an important organ for silk-secreting insects. Herein, we studied the effects of trace pyriproxyfen on autophagy and apoptosis of the silk gland in the lepidopteran model insect, Bombyx mori (silkworm). After treating fifth instar silkworm larvae with pyriproxyfen for 24 h, we found significant shrinkage, vacuolization, and fragmentation in the posterior silk gland (PSG). In addition, the results of autophagy-related genes of ATG8 and TUNEL assay also demonstrated that autophagy and apoptosis in the PSG of the silkworm was induced by pyriproxyfen. RNA-Seq results showed that pyriproxyfen treatment resulted in the activation of juvenile hormone signaling pathway genes and inhibition of 20-hydroxyecdysone (20E) signaling pathway genes. Among the 1808 significantly differentially expressed genes, 796 were upregulated and 1012 were downregulated. Among them, 30 genes were identified for autophagy-related signaling pathways, such as NOD-like receptor signaling pathway and mTOR signaling pathway, and 30 genes were identified for apoptosis-related signaling pathways, such as P53 signaling pathway and TNF signaling pathway. Further qRT-PCR and in vitro gland culture studies showed that the autophagy-related genes Atg5, Atg6, Atg12, Atg16 and the apoptosis-related genes Aif, Dronc, Dredd, and Caspase1 were responsive to the treatment of pyriproxyfen, with transcription levels up-regulated from 24 to 72 h. In addition, ATG5, ATG6, and Dronc genes had a more direct response to pyriproxyfen treatment. These results suggested that pyriproxyfen treatment could disrupt the hormone regulation in silkworms, promoting autophagy and apoptosis in the PSG. This study provides more evidence for the research on the damage of juvenile hormone analogues to non-target organisms or organs in the environment, and provides reference information for the scientific and rational use of juvenile hormone pesticides.

The role of flow in the self-assembly of dragline spider silk proteins.

Hydrodynamic flow in the spider duct induces conformational changes in dragline spider silk proteins (spidroins) and drives their assembly, but the underlying physical mechanisms are still elusive. Here we address this challenging multiscale problem with a complementary strategy of atomistic and coarse-grained molecular dynamics simulations with uniform flow. The conformational changes at the molecular level were analyzed for single-tethered spider silk peptides. Uniform flow leads to coiled-to-stretch transitions and pushes alanine residues into ß sheet and poly-proline II conformations. Coarse-grained simulations of the assembly process of multiple semi-flexible block copolymers using multi-particle collision dynamics reveal that the spidroins aggregate faster but into low-order assemblies when they are less extended. At medium-to-large peptide extensions (50%-80%), assembly slows down and becomes reversible with frequent association and dissociation events, whereas spidroin alignment increases and alanine repeats form ordered regions. Our work highlights the role of flow in guiding silk self-assembly into tough fibers by enhancing alignment and kinetic reversibility, a mechanism likely relevant also for other proteins whose function depends on hydrodynamic flow.

Impact of soil moisture stress during the silk emergence and grain-filling in maize.

Suboptimal soil moisture during the growing season often limits maize growth and yield. However, the growth stage-specific responses of maize to soil moisture regimes have not been thoroughly investigated. This study investigated the response of maize to five different soil moisture regimes, that are, 0.25, 0.20, 0.15, 0.10, and 0.05 m3 m-3 volumetric water content (VWC), during flowering and grain-filling stages. Sub-optimal soil moisture at the flowering and grain-filling stages reduced ear leaf stomatal conductance by 73 and 64%, respectively. An increase in stress severity caused significant reductions in ear leaf chlorophyll content and greenness-associated vegetation indices across growth stages. Fourteen days of soil moisture stress during flowering delayed silk emergence, reduced silk length (19%), and silk fresh weight (34%). Furthermore, sub-optimal soil moisture caused a significant reduction in both kernel number (53%) and weight (54%). Soil moisture stress at the flowering had a direct impact on kernel number and an indirect effect on kernel weight. During grain-filling, disruption of ear leaf physiology resulted in a 34% decrease in kernel weight and a 43% decrease in kernel number. Unlike grain-filling, treatments at the flowering significantly reduced kernel starch (3%) and increased protein by 29%. These findings suggest that developing reproductive stage stress-tolerant hybrids with improved resilience to soil moisture stress could help reduce the yield gap between irrigated and rainfed maize.

Microbial degradation mechanism of historical silk revealed by proteomics and metabolomics.

Archaeological silk undergoes destructive and irreversible changes during the natural process of decay. However, in-depth studies on the influence of this biological factor are still lacking. Here, a combination of proteomics and metabolomics is proposed for the first time to explore the interaction between bacteria and historical silk during biodegradation, which provides information on changes at the molecular level of proteins and bacterial metabolites. Morphological observation revealed biofilms produced by Stenotrophomonas maltophilia and Pseudomonas alcaligenes when cultured in the stationary phase and confirmed severe deterioration of silk. Proteomics showed that S. maltophilia had an unbiased effect on silk fibroin, indicating its ability to disrupt both heavy and light chains, as well as other proteins, while P. alcaligenes showed an affinity for more disordered proteins. Analysis of bacterial metabolites showed that overall activity reduction and significant accumulation of fatty acid and phenol metabolites occurred after silk addition, suggesting that the presence of silk may inhibit the activity of an individual strain. This study provides a new insight into the microbial degradation mechanism of archaeological silk.

Comparative seasonal analysis of Eri silkworm (Samia ricini Donovan) gut composition: implications for lignocellulose degradation.

Conversion of biomass such as lignocelluloses to an alternative energy source can contribute to sustainable development. Recently, biomass-degrading enzymes are reported to be common resources in insect-microbe interacting systems. Northeast India harbors ample sericigenous insect resources which are exploited for their silk products. Samia ricini Donovan is an economically important poly-phytophagous silkmoth capable of digesting foliage from different plant species, suggesting the versatility of a robust gut system. Here, a gut bacterial profile was determined by 16S rRNA gene characterization across the holometabolous life cycle during the summer and winter seasons, revealing 3 phyla, 13 families, and 22 genera. Comparative analysis among the seasonal gut isolates revealed a high diversity in summer, predominated by the genus Bacillus due to its high occurrence in all developmental stages. Shannon's diversity index demonstrated the second and fourth instars of summer as well as the fifth instar of winter to be relatively better developmental stages for gut bacteria assembly. Bacterial community shifts in concert to host developmental changes were found to be apparent between early instars and late instars in summer, which differed from those of winter. Forty-three and twenty-nine gut bacterial isolates were found to be cellulolytic and xylanolytic enzyme producers, respectively. The present results illustrate the gut microbiota of S. ricini over the seasons and support the holometabolous life cycle effect as the most likely factor shaping the gut bacterial microbiota. These findings may provide leads for the development of new cleaner and environmentally friendly lignocellulose-degrading enzymes.