BACKGROUND: Forest frog is a rare medicinal animal in China, but the skin of forest frog is waste after Oviductus Ranae production. The forest frog skin is rich of collagen, and is large in quantity without pollution and disease risk. So the forest frog skin has potential to be developed into collagen sponge; however, there is no research on collagen sponge preparation as yet.OBJECTIVE: To optimize the preparation of collagen sponge from forest frog skin, and to investigate the physical properties and in vitro cell compatibility.METHODS: Chitosan and glutaraldehyde were used to modify collagen sponge from forest frog skin. Chitosan/collagen (w/w) (1:1, 1:2, 1:4) and glutaraldehyde concentration (1%, 1.5% and 2%) were selected as the experimental factors.The significant water absorbency, mechanical properties and thermal denaturation temperature were chosen as the indexes. Using the orthogonal experimental design, we optimized collagen sponge preparation process. We also investigated the in vitro cell compatibility and surface morphology of the collagen sponge. The nine kinds of collagen sponges from forest frog skins were co-cultured with human foreskin fibroblasts to detect cell proliferation.RESULTS AND CONCLUSION: When the chitosan/collagen was 1:1 and the glutaraldehyde concentration was 1%, we could get the collagen sponge with ideal water absorbency (water absorption capacity=5.22), mechanical properties (elongation at break=10.96%) and thermal denaturation temperature (81.24 ℃). The aperture of the forest frog skin collagen sponge was 200-400 μm, and the pores were consistent in the size and arranged regularly. Except the chitosan/collagen of 1:4 and the glutaraldehyde concentration of 1% or 5%, all kinds of forest frog skin collagen sponges could promote the viability of human foreskin fibroblasts and exerted benefits to cell viability and growth. To conclude,the forest frog skin collagen sponge has good biocompatibility and apparent morphology, in aggreement with the requirements of biological materials.

This study aims to investigate the synergistic antimicrobial effect of demethylzeylasteral meropenem against NDM-1-positive Escherichia coli.Firstly,the nitrocefin hydrolysis assay and molecular docking assay were used to determine the inhibitory effect and mechanism of demethyl-zeylasteral on the hydrolytic activity of the NDM-1 protein.Secondly,the synergistic bacteriostatic effect of demethylzeylasteral with the carbapenem antibiotic meropenem was confirmed by the checkerboard minimum inhibitory concentration assay,growth curve assay,and inhibition zone as-say.The synergistic bactericidal effect of the combination was further determined by the time-kill-ing assay,live/dead bacterial staining assay,and membrane integrity assay.Finally,the G.mellonel-la infection model demonstrated the protective effect of demethylzeylasteral and meropenem.The results indicated that demethylzeylasteral significantly inhibited the hydrolytic activity of NDM-1(IC50=0.32 mg/L)and competitively affected the binding of NDM-1 to meropenem.In vitro tests showed that demethylzeylasteral had good a synergistic antibacterial effect with meropenem and could exert a synergistic bactericidal effect by enhancing the membrane damage of bacteria caused by meropenem.In vivo assays demonstrated that demethylzeylasteral increased the protective effect of meropenem from 67％to 80％.The results obtained in this study provide a therapeutic strategy and antibacterial synergist for treating NDM-1-positive bacterial infections.

Two types of Mycobacterium abscesses (Mab) were found in sputum from a patient with severe pneumonia in May 2020. One was Mycobacteriumabscessus with smooth morphology (Mab S), the other was Mycobacteriumabscessus with rough morphology (Mab R). Both of them were compared and drug susceptibility testing were performed to provide clinical scientific diagnosis and treatment. Morphology, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and 16S rRNA were used to analyze Mab S and Mab R, phylogenetic evolution tree was constructed by gene sequence alignment for homology, proportion and broth drug test were used for in vitro drug sensitivity test. There were morphological differences between Mab S and Mab R. MALDI-TOF MS analysis showed that there were 223 protein peaks in Mab R and 147 protein peaks in Mab S. Mab S contained 1 397 bp and Mab R contained 1 402 bp as 16s rRNA gene sequencing revealed. Drug susceptibility testing showed that both of them were almost resistant to all antituberculosis drugs, but sensitive to most of antibiotics. Mab S and Mab R were not only different in manifestations, but also in protein and gene comparison. Both of them were generally resistant to antituberculosis drugs. Antibiotic combined therapy has been confirmed to be an effective treatment in clinic.

Bivalves are species-rich mollusks with prominent protective roles in coastal ecosystems.Across these ancient lineages,colony-founding larvae anchor themselves either by byssus produc-tion or by cemented attachment.The latter mode of sessile life is strongly molded by left-right shell asymmetry during larval development of Ostreoida oysters such as Crassostrea hongkongensis.Here,we sequenced the genome of C.hongkongensis in high resolution and compared it to reference bivalve genomes to unveil genomic determinants driving cemented attachment and shell asymmetry.Importantly,loss of the homeobox gene Antennapedia(Antp)and broad expansion of lineage-specific extracellular gene families are implicated in a shift from byssal to cemented attachment in bivalves.Comparative transcriptomic analysis shows a conspicuous divergence between left-right asymmetrical C.hongkongensis and symmetrical Pinctada fucata in their expression profiles.Especially,a couple of orthologous transcription factor genes and lineage-specific shell-related gene families including that encoding tyrosinases are elevated,and may cooperatively govern asymmet-rical shell formation in Ostreoida oysters.