Microbial Biotechnology最新文献

The transcriptional architecture of phages can deepen our understanding of the phage-host infection process and can be of key importance for phage engineering and biotechnological applications. Here, we applied ONT-cappable-sequencing, a long-read RNA-sequencing technique, to study the regulatory mechanisms of Pseudomonas infecting giant phage 201ϕ2-1. We identified 67 promoters and 132 terminators that together represent 92 transcriptional units. A full comparison of these data to the transcriptome of model Pseudomonas phage ϕKZ confirmed that the transcriptional programs of these prototypes of the Serwervirus and Phikzvirus genera are largely conserved, despite some subtle regulatory differences. Evidence supporting these shared mechanisms include the identification of highly similar sequence motifs for regulatory elements in both phages and the conservation of regulatory elements loci relative to homologous genes in each phage. Moreover, we discovered a sRNA in 201ϕ2-1 that is highly conserved among prototype members of different giant phage genera. Sequencing of the 201ϕ2-1 host genome resulted in its reclassification as Pseudomonas atacamensis, a close relative of the important agricultural biocontrol agent Pseudomonas chlororaphis. Finally, we conducted in vivo assays of eight 201ϕ2-1 terminators and found them to strongly terminate transcription in P. chlororaphis. Control elements from phage transcriptional programs have a rich history for applications in biotechnology. In these studies, we demonstrate new insight into the transcriptional program of 201ϕ2-1 and demonstrate the potential of its regulatory elements for novel and useful tools for synthetic biology circuitry.

Merkel cell polyomavirus (MCPyV) is the foremost causative factor of Merkel cell carcinoma (MCC), a rare yet highly aggressive skin cancer. Although the evaluation of circulating IgG antibodies against Merkel cell polyomavirus (MCPyV) LT/sT oncoproteins is clinically useful for MCC diagnosis/prognosis, a limited number of assays for identifying such antibodies have been developed. Herein, a novel indirect immunoassay with synthetic epitopes/mimotopes of MCPyV oncoproteins was computationally designed and experimentally validated on control sera and sera from healthy individuals and MCC patients. Upon computational design of five synthetic peptides, the performance of the immunoassay in detecting anti-oncoprotein IgGs in MCPyV-positive and -negative control sera was evaluated. The immunoassay was afterwards extended on sera from healthy individuals, and, for longitudinal analysis, MCC patients. Performance properties such as sensitivity and specificity and positive/negative predictive values were adequate. Receiver-operating characteristic (ROC) curves indicated that the areas under the curves (AUCs) were within the low/moderately accurate ranges. Immunoassay was repeatable, reproducible and accurate. As expected, the serum anti-oncoprotein IgG prevalence in healthy individuals was low (2%–5%). Anti-oncoprotein IgGs slightly increased when MCC patients experienced partial tumour remission and/or stable disease, compared to baseline. Our data indicate that the newly developed immunoassay is reliable for detecting circulating anti-oncoprotein IgGs both in healthy individuals and MCC patients.

A healthy mucus is essential for maintaining intestinal homeostasis and overall well-being. In recent years, extensive research focused on understanding the intricate interactions between mucus and the gut microbiota. Mucus-adhering bacteria play crucial roles in preserving barrier integrity, epithelial permeability and mucus architecture, as well as in the colonization resistance against pathogens. Unravelling the significance of these microorganisms in human health and disease is challenging, primarily because most of the studies on the human gut microbiota rely on faecal samples, which do not fully represent the microecological complexity found in the intestinal mucosa. This review discusses novel strategies to specifically target and evaluate the mucosal microbiota, such as culturomics applied to mucosal biopsies or brushings, intestinal organoids and artificial in vitro models incorporating mucus.

Bacterial infections represent a significant global threat to human health, leading to considerable economic losses through increased healthcare costs and reduced productivity. One major challenge in treating these infections is the presence of biofilms - structured bacterial communities that form protective barriers, making traditional treatments less effective. Additionally, the rise of antibiotic-resistant bacteria has exacerbated treatment difficulties. To address these challenges, researchers are developing and exploring innovative approaches to combat biofilm-related infections. This mini-review highlights recent advancements in the following key areas: surface anti-adhesion technologies, electricity, photo/acoustic-active materials, endogenous mimicking agents, and innovative drug delivery systems. These strategies aim to prevent biofilm formation, disrupt existing biofilms, and enhance the efficacy of antimicrobial treatments. Currently, these approaches show great potential for applications in medical fields such as medical device and wound – associated biofilm infections. By summarizing these developments, this mini-review provides a comprehensive resource for researchers seeking to advance the management and treatment of biofilm-associated infections.

Previous reports have demonstrated that alcohol consumption significantly reduces the abundance of Lactobacillus in the gut. In this study, we selected five species of the genus Lactobacillus, commonly found in fermented foods, and acknowledged them as safe, edible, and effective in preventing or treating certain diseases, to evaluate their effects on alcoholic liver disease (ALD). By comparing the liver damage indices in each group, we found that the type strain of Lactobacillus helveticus (LH, ATCC 15009) had the most marked alleviating effect on ALD-induced liver injury. Furthermore, experiments combining microbiomics and metabolomics were conducted to explore the mechanisms underlying the hepatoprotective effects of LH. Finally, we discovered that LH mitigated ethanol-induced liver steatosis and inflammation in ALD mice by altering the structure and function of the gut microbiome, increasing intestinal levels of short-chain fatty acids (SCFAs), and enhancing gut barrier integrity. These findings suggest a potential strategy for the clinical management of patients with ALD.

Infected wounds can result in complex clinical complications and delayed healing, presenting a significant global public health challenge. This study explored the effects of topical application of two probiotics, Lactobacillus rhamnosus GG (LGG) and Bifidobacterium animalis subsp. lactis BB-12, on the microenvironment of infected wounds and their impact on wound healing. LGG and BB-12 were applied separately and topically on the Staphylococcus aureus (S. aureus)-infected skin wounds of the rat model on a daily basis. Both probiotics significantly accelerated wound healing, demonstrated by enhanced granulation tissue formation and increased collagen deposition, with BB-12 showing superior efficacy. LGG and BB-12 both effectively inhibited neutrophil infiltration and decreased the expression of pro-inflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). Notably, BB-12 markedly reduced IL-6 levels, while LGG significantly lowered TNF-α, transforming growth factor-β (TGF-β) and vascular endothelial growth factor (VEGF). Additionally, both probiotics promoted macrophage polarization towards the anti-inflammatory M2 phenotype. Microbiota analysis revealed that LGG and BB-12 significantly decreased the abundance of pathogenic bacteria (e.g. Staphylococcus and Proteus) and increased the proportion of beneficial bacteria (e.g. Corynebacterium). Particularly, BB-12 was more effective in reducing Staphylococcus abundance, whereas LGG excelled in promoting Corynebacterium growth. These findings suggest the ability of LGG and BB-12 to modulate the wound microenvironment, enhance wound healing and provide valuable insights for the management of infected wounds.

In situ vaccination is a therapeutic approach aimed at exploiting tumour antigens available at a tumour site to induce tumour-specific adaptive immune responses. Antigens released from dying tumour cells are assumed to be taken up by activated dendritic cells and presented to T cells that seek out and destroy tumour cells. This process is significantly impeded in the immunosuppressive microenvironment of tumours. There is a growing trend in in situ vaccine strategies that utilize bacteria as natural adjuvants or as factories for cytokines, aiming to enhance the presentation of in situ antigens by antigen-presenting cells. Recently, a novel approach using flagellate bacteria-mediated antigen delivery to activate dendritic cells has been proposed. This method actively facilitates the delivery of intratumoral antigens, improving their presentation for in situ cancer vaccination. Here, we highlight how flagellate bacteria-mediated antigen delivery enhances the immune activation capabilities of in situ vaccines. Meanwhile, we provide perspectives and outlooks on these promising antigen delivery technologies.

Fusarium wilt is one of the major constraints on global watermelon production, and Fusarium oxysporum f. sp. niveum (Fon) is the causative agent of Fusarium wilt in watermelon and results in severe yield and quality losses worldwide. The enhancement of antifungal activity from antagonistic bacteria against Fon is highly practical for managing Fusarium wilt in watermelon. The aim of this study was to maximize the antifungal activity of Bacillus velezensis LZN01 by optimizing fermentation conditions and analysing its regulatory mechanism via transcriptome sequencing. The culture and fermentation conditions for strain LZN01 were optimized by single-factor and response surface experiments. The optimum culture conditions for this strain were as follows: the addition of D-fructose at 35 g/L and NH₄Cl at 5 g/L in LB medium, pH 7, and incubation at 30°C for 72 h. The fungal inhibition rate for strain LZN01 reached 71.1%. The improvement of inhibition rate for strain LZN01 in optimization fermentation was supported by transcriptomic analysis; a total of 491 genes were upregulated, while 736 genes were downregulated. Transcriptome analysis revealed that some differentially expressed genes involved in carbon and nitrogen metabolism, oxidation–reduction, fatty acid and secondary metabolism; This optimization process could potentially lead to significant alterations in the production levels and types of antimicrobial compounds by the strain. Metabolomics and UPLC/Q-Exactive Orbitrap MS analysis revealed that the production yields of antimicrobial compounds, such as surfactin, fengycin, shikimic acid, and myriocin, increased or were detected in the cell-free supernatant (CFS) after the fermentation optimization process. Our results indicate that fermentation optimization enhances the antifungal activity of the LZN01 strain by influencing the expression of genes responsible for the synthesis of antimicrobial compounds.

Bacterial surface display in combination with fluorescence-activated cell sorting is a versatile and robust system and an interesting alternative approach to phage display for the generation of therapeutic affinity proteins. The system enables real-time monitoring and sorting of cell populations, which presents unique possibilities for drug development. It has been used to develop several affibody molecules currently being evaluated preclinically for the treatment and diagnosis of, for example, cancer and neurodegenerative diseases. Additionally, it can be implemented in other areas of drug design, such as for mapping epitopes and evolving enzyme specificities.

Escherichia coli (E. coli) is a ubiquitous symbiotic bacterium in the gut, and the diversity of E. coli genes determines the diversity of its functions. In this review, the two-edged sword theory was innovatively proposed. For the question ‘how can we harness the ambivalent nature of E. coli to screen and treat CRC?’, in terms of CRC screening, the variations in the abundance and subtypes of E. coli across different populations present an opportunity to utilise it as a biomarker, while in terms of CRC treatment, the natural beneficial effect of E. coli on CRC may be limited, and engineered E. coli, particularly certain subtypes with probiotic potential, can indeed play a significant role in CRC treatment. It seems that the favourable role of E. coli as a genetic tool lies not in its direct impact on CRC but its potential as a research platform that can be integrated with various technologies such as nanoparticles, imaging methods, and synthetic biology modification. The relationship between gut microflora and CRC remains unclear due to the complex diversity and interaction of gut microflora. Therefore, the application of E. coli should be based on the ‘One Health’ view and take the interactions between E. coli and other microorganisms, host, and environmental factors, as well as its own changes into account. In this paper, the two-edged sword role of E. coli in CRC is emphasised to realise the great potential of E. coli in CRC screening and treatment.