Recently, the drawbacks arising from the overuse of antibiotics have drawn growing public attention. Among them, drug-resistance (DR) and even multidrug-resistance (MDR) pose significant challenges in clinical practice. As a representative of a DR or MDR pathogen, Staphylococcus aureus can cause diversity of infections related to different organs, and can survive or adapt to the diverse hostile environments by switching into other phenotypes, including biofilm and small colony variants (SCVs), with altered physiologic or metabolic characteristics. In this review, we briefly describe the development of the DR/MDR as well as the classical mechanisms (accumulation of the resistant genes). Moreover, we use multidimensional scaling analysis to evaluate the MDR relevant hotspots in the recent published reports. Furthermore, we mainly focus on the possible non-classical resistance mechanisms triggered by the two important alternative phenotypes of the S. aureus, biofilm and SCVs, which are fundamentally caused by the different global regulation of the S. aureus population, such as the main quorum-sensing (QS) and agr system and its coordinated regulated factors, such as the SarA family proteins and the alternative sigma factor σB (SigB). Both the biofilm and the SCVs are able to escape from the host immune response, and resist the therapeutic effects of antibiotics through the physical or the biological barriers, and become less sensitive to some antibiotics by the dormant state with the limited metabolisms.

In this study, the yield of exopolysaccharide (EPS) from Lactobacillus plantarum R301 was optimized using a single-factor experiment and response surface methodology (RSM). After optimization, the EPS yield was increased with a fold-change of 0.85. The significant factors affecting EPS production, as determined through a Plackett–Burman design and Central Composite Design (CCD), were MgSO₄ concentration, initial pH, and inoculation size. The maximum yield was 97.85 mg/mL under the condition of 0.01% MgSO₄, an initial pH 7.4, and 6.4% of the inoculation size. In addition, the EPS exhibited strong antioxidant activity, as demonstrated by its ability to scavenge DPPH, ABTS, and hydroxyl radicals. The scavenging rate was up to 100% at concentrations of 4 mg/mL, 1 mg/mL, and 2 mg/mL, respectively. Moreover, the EPS also exhibited reducing power, which was about 30% that of ascorbic acid when both tended to be stable with the increased concentration. These results suggest that L. plantarum R301 EPS possesses different antioxidant mechanisms and warrants further investigation. In addition to its antioxidant activity, the EPS also demonstrated good anti-inflammatory activity by inhibiting the inflammation induced by lipopolysaccharide (LPS) in RAW 264.7 cells, which could decrease nitric oxide (NO) production and expression of the proinflammatory cytokine Il-6. These findings suggest that L. plantarum R301 EPS could be used as a potential multifunctional food additive in the food industry.

The hybrid peptide cecropin A (1–8)–LL37 (17–30) (C–L), derived from the sequence of cecropin A (C) and LL-37 (L), showed significantly increased antibacterial activity and minimized hemolytic activity than C and L alone. To obtain high-level production of C–L, the deoxyribonucleic acid sequence encoding C–L with preferred codons was cloned into pET-SUMO to construct a fusion expression vector, and overexpressed in Escherichia coli (E. coli) BL21 (DE3). The maximum fusion protein (92% purity) was obtained with the yield of 89.14 mg/L fermentation culture after purification with Ni-NTA Sepharose column. The hybrid C–L was cleaved from the fusion protein by SUMO-protease, and 17.54 mg/L pure active C–L was obtained. Furthermore, the purified C–L showed identical antibacterial and hemolytic activity to synthesized C–L. Stability analysis results exhibited that the activity of C–L changed little below 80 °C for 20 min, but when the temperature exceeded 80 °C, a significant decrease was observed. Varying the pH from 5.0 to 10.0 did not appear to influence the activity of C–L, however, pH below 4.0 decreased the antibacterial activity of C–L rapidly. Under the challenge of several proteases (pepsin, trypsin, and proteinase K), the functional activity of C–L was maintained over 50%. In summary, this study not only supplied an effective approach for high-level production of hybrid peptide C–L, but paved the way for its further exploration in controlling infectious diseases of farm animals or even humans.

This study pertains to the new approach for the development of hybrid peptide LL-37Tα1 and its biomedical applications. A linear cationic hybrid peptide, LL-37Tα1 was derived from two parental peptides (LL-37 and Tα1) recognized as potent anti-endotoxin without any hemolytic or cytotoxic activity. We successfully cloned the gene of hybrid peptide LL-37Tα1 in PpICZαA vector and expressed in the Pichia pastoris. The recombinant peptide was purified by Ni-affinity column and reverse-phase high performance liquid chromatography (RP-HPLC) with an estimated molecular mass of 3.9 kDa as determined by SDS-PAGE and mass spectrometry. We analyzed the LPS neutralization by limulus amebocyte lysate (LAL) activity and the results indicate that the hybrid peptide LL-37Tα1 directly binds endotoxin and significantly (p < 0.05) neutralizes the effect of LPS in a dose-dependent manner. Lactate dehydrogenase (LDH) assay revealed that LL-37Tα1 successfully reduces the LPS-induced cytotoxicity in mouse RAW264.7 macrophages. Moreover, it significantly (p < 0.05) decreased the levels of nitric oxide, proinflammatory cytokines including TNF-α, IL-6, IL-1β, and diminished the number of apoptotic cells in LPS-stimulated mouse RAW264.7 macrophages. Our results suggest that the P. pastoris expression system is cost-effective for commercial production of the immunomodulatory and anti-inflammatory hybrid peptide (IAHP) LL-37Tα1 and the peptide may serve as effective anti-endotoxin/anti-inflammatory agent with minimal cytotoxicity.

Mulberry is a rapidly growing plant that thrives in diverse climatic, topographical, and soil types, spanning temperature and temperate countries. Mulberry plants are valued as functional foods for their abundant chemical composition, serving as a significant reservoir of bioactive compounds like proteins, polysaccharides, phenolics, and flavonoids. Moreover, these compounds displayed potent antioxidant activity by scavenging free radicals, inhibiting reactive oxygen species generation, and restoring elevated nitric oxide production induced by LPS stimulation through the downregulation of inducible NO synthase expression. Active components like oxyresveratrol found in Morus demonstrated anti-inflammatory effects by inhibiting leukocyte migration through the MEK/ERK signaling pathway. Gallic and chlorogenic acids in mulberry leaves (ML) powder-modulated TNF, IL-6, and IRS1 proteins, improving various inflammatory conditions by immune system modulation. As we delve deeper into understanding its anti-inflammatory potential and how it works therapeutically, it is crucial to refine the extraction process to enhance the effectiveness of its bioactive elements. Recent advancements in extraction techniques, such as solid–liquid extraction, pressurized liquid extraction, superficial fluid extraction, microwave-assisted extraction, and ultrasonic-assisted extraction, are being explored. Among the extraction methods tested, including Soxhlet extraction, maceration, and ultrasound-assisted extraction (UAE), UAE demonstrated superior efficiency in extracting bioactive compounds from mulberry leaves. Overall, this comprehensive review sheds light on the potential of mulberry as a natural immunomodulatory agent and provides insights into its mechanisms of action for future research and therapeutic applications.

EF-1 is a novel peptide derived from two bacteriocins, plantaricin E and plantaricin F. It has a strong antibacterial activity against Escherichia coli and with negligible hemolytic effect on red blood cells. However, the chemical synthesis of EF-1 is limited by its high cost. In this study, we established a heterologous expression of EF-1 in Pichia pastoris. The transgenic strain successfully expressed hybrid EF-1 peptide, which had a molecular weight of ~5 kDa as expected. The recombinant EF-1 was purified by Ni²⁺ affinity chromatography and reversed-phase high performance liquid chromatography (RP-HPLC), which achieved a yield of 32.65 mg/L with a purity of 94.9%. The purified EF-1 exhibited strong antimicrobial and bactericidal activities against both Gram-positive and -negative bacteria. Furthermore, propidium iodide staining and scanning electron microscopy revealed that EF-1 can directly induce cell membrane permeabilization of E. coli. Therefore, the hybrid EF-1 not only preserves the individual properties of the parent peptides, but also acquires the ability to disrupt Gram-negative bacterial membrane. Meanwhile, such an expression system can reduce both the time and cost for large-scale peptide production, which ensures its potential application at the industrial level.