Synthesis of di-rhamnolipids by the avirulent, mono-rhamnolipid producing strain Pseudomonas aeruginosa ATCC 9027.

To construct a derivative of the avirulent Pseudomonas aeruginosa ATCC 9027 that produces high levels of di-rhamnolipid, that has better physico-chemical characteristics for biotechnological applications than mono-rhamnolipid, which is the sole type produced by ATCC 9027. We used plasmids expressing the rhlC gene, which encodes for rhamnosyl transferase II that transforms mono- to di-rhamnolipids under different promoters and in combination with the gene coding for the RhlR quorum sensing regulator, or the mono-rhamnolipid biosynthetic rhlAB operon. The plasmids tested carrying the rhlC gene under the lac promoter were plasmid prhlC and prhlRC, while prhlAB-R-C expressed this gene from the rhlA promoter, forming part of the artificially constructed rhlAB-R-C operon. We measured rhamnolipds concentrations using the orcinol method and determined the proportion of mono-rhamnolipids and di-rhamnolipids by UPLC/MS/MS. We found that the expression of rhlC in P. aeruginosa ATCC 9027 caused the production of di-rhamnolipids and that the derivative carrying plasmid prhlAB-R-C gives the best results considering total rhamnolipids and a higher proportion of di-rhamnolipids. A P. aeruginosa ATCC 9027 derivative with increased di-rhamnolipids production was developed by expressing plasmid prhlAB-R-C, that produces similar rhamnolipids levels as PAO1 type-strain and presented a higher proportion of di-rhamnolipids than this type-strain.

The biological activity of bacterial rhamnolipids on Arabidopsis thaliana and the cyst nematode Heterodera schachtii is linked to their molecular structure.

Rhamnolipids (RLs) are amphiphilic compounds of bacterial origin that offer a broad range of potential applications as biosurfactants in industry and agriculture. They are reported to be active against different plant pests and pathogens and thus are considered promising candidates for nature-derived plant protection agents. However, as these glycolipids are structurally diverse, little is known about their exact mode of action and, in particular, the relation between molecular structure and biological activity against plant pests and pathogens. Engineering the synthesis pathway in recombinant Pseudomonas putida strains in combination with advanced HPLC techniques allowed us to separately analyze the activities of mixtures of pure mono-RLs (mRLs) and of pure di-RL (dRLs), as well as the activity of single congeners. In a model system with the plant Arabidopsis thaliana and the plant-parasitic nematode (PPN) Heterodera schachtii we demonstrate that RLs can significantly reduce infection, whereas their impact on the host plant varied depending on their molecular structure. While mRLs reduced plant growth even at a low concentration, dRLs showed a neutral to beneficial impact on plant development. Treating plants with dRLs triggered an increased reactive oxygen species (ROS) production, indicating the activation of stress-response signaling and possibly plant defense. Pretreatment of plants with mRLs or dRLs prior to application of flagellin (flg22), a known ROS inducer, further increased the ROS response to flg22. While dRLs stimulated an elevated flg22-induced ROS peak, a pretreatment with mRLs resulted in a prolonged synthesis of ROS indicating a generally elevated stress level. Neither mRLs nor dRLs induced the expression of plant defense marker genes of salicylic acid, jasmonic acid, and ethylene pathways. Detailed studies on dRLs revealed that even high concentrations up to 755 ppm of these molecules have no lethal impact on H. schachtii infective juveniles. Infection assays with individual dRL congeners showed that the C10-C8 acyl chained dRL was the only congener without effect, while dRLs with C10-C12 and C10-C12:1 acyl chains were most efficient in reducing nematode infection even at concentrations below 2 ppm. As determined by phenotyping and ROS measurements, A. thaliana reacted more sensitive to long-chained dRLs in a concentration-dependent manner. Our experiments show a clear structure-activity relation for the effect of RLs on plants. In conclusion, functional assessment and analysis of the mode of action of RLs in plants and other organisms require careful consideration of their molecular structure and composition.

Exploration and characterization of a newly isolated bacterium, Enterobacter quasihormaechei strain BDIFST24001, capable of producing rhamnolipid biosurfactant for oil remediation.

Biosurfactants are naturally occurring compounds synthesized by micro-organisms that increasingly attract attention due to both their living area and application in various industries. In this study, we explore and characterize a novel bacterium, Enterobacter quasihormaechei strain BDIFST24001, isolated for its ability to produce rhamnolipid biosurfactants, with the aim of facilitating oil remediation processes. The isolation of this bacterium was carried out using Luria-Bertani (LB) broth media from environmental samples collected from oil-contaminated sites in Dhaka City. Screening tests, including the oil spreading method and drop collapse assay, were conducted to identify potential biosurfactant-producing strains, leading to the selection of E. quasihormaechei strain BDIFST24001 based on its favourable performance. Subsequent molecular identification revealed a high similarity of the strain's 16S rRNA gene to E. quasihormaechei, which was corroborated through phylogenetic analysis. Further analysis of the biosurfactant produced by this strain indicated its rhamnolipid nature, as confirmed by FT-IR spectroscopy. The rhamnolipids exhibited promising surface-active properties, including a significant reduction in surface tension and emulsification activity, as evidenced by surface tension measurements and emulsification index assays. Optimization studies revealed that the optimal conditions for rhamnolipid production by E. quasihormaechei strain BDIFST24001 were a temperature of 37 °C, pH 10.0 and salinity of 4 %. The rhamnolipids produced by this strain demonstrated effective oil remediation capabilities, as observed through controlled experiments using petrol oil. The rhamnolipids effectively reduced the surface tension of the oil-water interface, facilitating the dispersion and emulsification of the oil phase in water. Overall, our findings highlight the potential of E. quasihormaechei strain BDIFST24001 as a promising candidate for biosurfactant-mediated oil spill cleanup and environmental remediation efforts.

Antimicrobial activity, foaming properties, and interacting mechanism of rhamnolipids in presence of silk fibroin through spectroscopy, molecular docking, and microbiological experiments.

As a type of biosurfactant, rhamnolipids (RLs) are multifunctional skin-care ingredients, and the molecular interaction of RLs with silk fibroin (SF) is a more complicated process than has long been believed. The interaction and functional properties of them, and their potential as fungicidal agents for agricultural products and as organic preservatives for cosmetics were assessed in this paper. The SF addition makes the RLs aggregation easier through the complexes formation, which decreases the applied concentration of surfactant. The results of spectroscopic analyses and molecular docking suggest that hydrogen bonding and van der Waals forces are significant contributed to the binding mechanism between the two substances. The addition of SF notably enhances the foaming capacity and stability of RLs. The certain antibacterial and antifungal properties of RLs are basically not affected by the SF addition, even the SF-RLS system demonstrates an unobvious synergistic inhibitory impact on Glomerella cingulate (GC). The results offer a theoretical framework for the utilization of RLs as natural fungicides and preservatives in presence of nutritional components, considering the properties of RLs as nontoxic, biodegradable, environmentally friendly, and good compatibility.

Rhamnolipids and fengycins interact differently with biomimetic lipid membrane models of Botrytis cinerea and Sclerotinia sclerotiorum: Lipidomics profiles and biophysical studies.

Rhamnolipids (RLs) and Fengycins (FGs) are biosurfactants with very promising antifungal properties proposed to reduce the use of synthetic pesticides in crops. They are amphiphilic molecules, both known to target the plasma membrane. They act differently on Botrytis cinerea and Sclerotinia sclerotiorum, two close Sclerotiniaceae phytopathogenic fungi. RLs are more efficient at permeabilizing S. sclerotiorum, and FGs are more efficient at permeabilizing B. cinerea mycelial cells. To study the link between the lipid membrane composition and the activity of RLs and FGs, we analyzed the lipid profiles of B. cinerea and S. sclerotiorum. We determined that unsaturated or saturated C18 and saturated C16 fatty acids are predominant in both fungi. We also showed that phosphatidylethanolamine (PE), phosphatidic acid (PA), and phosphatidylcholine (PC) are the main phospholipids (in this order) in both fungi, with more PA and less PC in S. sclerotiorum. The results were used to build biomimetic lipid membrane models of B. cinerea and S. sclerotiorum for all-atom molecular dynamic simulations and solid-state NMR experiments to more deeply study the interactions between RLs or FGs with different compositions of lipid bilayers. Distinctive effects are exerted by both compounds. RLs completely insert in all the studied model membranes with a fluidification effect. FGs tend to form aggregates out of the bilayer and insert individually more easily into the models representative of B. cinerea than those of S. sclerotiorum, with a higher fluidification effect. These results provide new insights into the lipid composition of closely related fungi and its impact on the mode of action of very promising membranotropic antifungal molecules for agricultural applications.

SkinCom, a synthetic skin microbial community, enables reproducible investigations of the human skin microbiome.

Existing models of the human skin have aided our understanding of skin health and disease. However, they currently lack a microbial component, despite microbes' demonstrated connections to various skin diseases. Here, we present a robust, standardized model of the skin microbial community (SkinCom) to support in vitro and in vivo investigations. Our methods lead to the formation of an accurate, reproducible, and diverse community of aerobic and anaerobic bacteria. Subsequent testing of SkinCom on the dorsal skin of mice allowed for DNA and RNA recovery from both the applied SkinCom and the dorsal skin, highlighting its practicality for in vivo studies and -omics analyses. Furthermore, 66% of the responses to common cosmetic chemicals in vitro were in agreement with a human trial. Therefore, SkinCom represents a valuable, standardized tool for investigating microbe-metabolite interactions and facilitates the experimental design of in vivo studies targeting host-microbe relationships.

Bioactive baicalin rhamno-nanocapsules as phytotherapeutic platform for treatment of acute myeloid leukemia.

Leukemia, particularly acute myeloid leukemia (AML) is considered a serious health condition with high prevalence among adults. Accordingly, finding new therapeutic modalities for AML is urgently needed. This study aimed to develop a biocompatible nanoformulation for effective oral delivery of the phytomedicine; baicalin (BAC) for AML treatment. Lipid nanocapsules (LNCs) based on bioactive natural components; rhamnolipids (RL) as a biosurfactant and the essential oil linalool (LIN), were prepared using a simple phase-inversion method. The elaborated BAC-LNCs displayed 61.1 nm diameter and 0.2 PDI. Entrapment efficiency exceeded 98 % with slow drug release and high storage-stability over 3 months. Moreover, BAC-LNCs enhanced BAC oral bioavailability by 2.3-fold compared to BAC suspension in rats with higher half-life and mean residence-time. In vitro anticancer studies confirmed the prominent cytotoxicity of BAC-LNCs on the human leukemia monocytes (THP-1). BAC-LNCs exerted higher cellular association, apoptotic capability and antiproliferative activity with DNA synthesis-phase arrest. Finally, a mechanistic study performed through evaluation of various tumor biomarkers revealed that BAC-LNCs downregulated the angiogenic marker, vascular endothelial growth-factor (VEGF) and the anti-apoptotic marker (BCl-2) and upregulated the apoptotic markers (Caspase-3 and BAX). The improved efficacy of BAC bioactive-LNCs substantially recommends their pharmacotherapeutic potential as a promising nanoplatform for AML treatment.

Rhamnolipids stabilized essential oils microemulsion for antimicrobial and fruit preservation.

Essential oils, well-known for their antifungal properties, are widely utilized to combat fruit decay. However, their application faces big challenges due to their high volatility and hydrophobic traits, which leads to strong odor, short effective time and poor dispersivity. This study aimed to address these challenges by formulating microemulsions consisting of essential oils and rhamnolipids. The optimized microemulsion, featuring a small particle size of 6.8 nm, exhibited higher stability and lower volatility than conventional emulsion. Notably, the prepared microemulsions demonstrated remarkable antimicrobial efficacy against E. coli, S. aureus, C. albicans, S. cerevisiae, and A. niger. The application of these microemulsions proved to be highly effective in preventing blueberry decay while preserving fruit's quality, particularly by minimizing the loss of essential nutrients such as anthocyanins. Consequently, essential oil microemulsions emerge as a highly effective postharvest preservative for fruits, offering a promising solution to extend their shelf life and enhance overall quality.

Antibacterial mechanism of Pseudomonas aeruginosa UKMP14T rhamnolipids against multidrug resistant Acinetobacter baumannii.

Rhamnolipids, a major category of glycolipid biosurfactant, have recently gained enormous attention in medical field because of their relevance as effective antibacterial agents against a wide variety of pathogenic bacteria. Our previous studies have shown that rhamnolipids from an environmental isolate of Pseudomonas aeruginosa UKMP14T possess antibacterial, anti-adhesive and anti-biofilm activity against multidrug-resistant ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacter sp.) pathogens. However, the mechanism of their antibacterial action remains unclear. Thus, this study aimed to elucidate the mechanism of the antibacterial action of P. aeruginosa UKMP14T rhamnolipids by studying the changes in cells of one of the ESKAPE pathogens, Acinetobacter baumannii, which is the most difficult strain to kill. Results revealed that rhamnolipid treatment rendered A. baumannii cells more hydrophobic as evaluated through contact angle measurements. It also induced the release of cellular proteins measuring 510 µg/mL at a rhamnolipid concentration of 1000 µg/mL. In addition, rhamnolipids were found to be bactericidal in their action as they could permeate the inner membranes, leading to a leak-out of nucleotides. More than 50 % of the cells were found to be killed upon 1000 µg/mL rhamnolipid treatment as observed through fluorescence microscopy. Other cellular changes such as irregular shape and size, membrane perturbations, clumping, shrinkage and physical damage were clearly visible in SEM, FESEM and laser micrographs. Furthermore, rhamnolipid treatment inhibited the levels of acyl-homoserine lactones (AHLs) in A. baumannii, which are vital for their biofilm formation and virulence. The obtained results indicate that P. aeruginosa UKMP14T rhamnolipids target outer and inner bacterial membranes through permeation, including physical damage to the cells, leading to cell leakage. Furthermore, AHL inhibition appears to be the mechanism behind their anti-biofilm action. All these observations can be correlated to rhamnolipids' antibacterial effect against A. baumannii.

Bioelectrochemical synthesis of rhamnolipids and energy production and its correlation with nitrogen in air-cathode microbial fuel cells.

Microbial fuel cells (MFCs) have been recently proven to synthesise biosurfactants from waste products. In classic bioreactors, the efficiency of biosynthesis process can be controlled by the concentration of nitrogen content in the electrolyte. However, it was not known whether a similar control mechanism could be applied in current-generating conditions. In this work, the effect of nitrogen concentration on biosurfactant production from waste cooking oil was investigated. The concentration of NH4Cl in the electrolyte ranged from 0 to 1 g L-1. The maximum power density equal to 17.5 W m-3 was achieved at a concentration of 0.5 g L-1 (C/N = 2.32) and was accompanied by the highest surface tension decrease (to 54.6 mN m-1) and an emulsification activity index of 95.4%. Characterisation of the biosurfactants produced by the LC-MS/MS method showed the presence of eleven compounds belonging to the mono- and di-rhamnolipids group, most likely produced by P. aeruginosa, which was the most abundant (19.6%) in the community. Importantly, we have found a strong correlation (R = -0.96) of power and biosurfactant activity in response to C/N ratio. This study shows that nitrogen plays an important role in the current-generating metabolism of waste cooking oil. To the best of our knowledge, this is the first study where the nitrogen optimisation was investigated to improve the synthesis of biosurfactants and power generation in a bioelectrochemical system.

Biological machinery for the production of biosurfactant and their potential applications.

The growing biotechnology industry has focused a lot of attention on biosurfactants because of several advantages over synthetic surfactants. These benefits include worldwide public health, environmental sustainability, and the increasing demand from sectors for environmentally friendly products. Replacement with biosurfactants can reduce upto 8% lifetime CO2 emissions avoiding about 1.5 million tons of greenhouse gas released into the atmosphere. Therefore, the demand for biosurfactants has risen sharply occupying about 10% (∼10 million tons/year) of the world production of surfactants. Biosurfactants' distinct amphipathic structure, which is made up of both hydrophilic and hydrophobic components, enables these molecules to perform essential functions in emulsification, foam formation, detergency, and oil dispersion-all of which are highly valued characteristic in a variety of sectors. Today, a variety of biosurfactants are manufactured on a commercial scale for use in the food, petroleum, and agricultural industries, as well as the pharmaceutical and cosmetic industries. We provide a thorough analysis of the body of knowledge on microbial biosurfactants that has been gained over time in this research. We also discuss the benefits and obstacles that need to be overcome for the effective development and use of biosurfactants, as well as their present and future industrial uses.

Unveiling the capacity of bioaugmentation application, in comparison with biochar and rhamnolipid for TPHs degradation in aged hydrocarbons polluted soil.

Persistent, aged hydrocarbons in soil hinder remediation, posing a significant environmental threat. While bioremediation offers an environmentally friendly and cost-effective approach, its efficacy for complex contaminants relies on enhancing pollutant bioavailability. This study explores the potential of immobilized bacterial consortia combined with biochar and rhamnolipids to accelerate bioremediation of aged total petroleum hydrocarbon (TPH)-contaminated soil. Previous research indicates that biochar and biosurfactants can increase bioremediation rates, while mixed consortia offer sequential degradation and higher hydrocarbon mineralization. The present investigation aimed to assess whether combining these strategies could further enhance degradation in aged, complex soil matrices. The bioaugmentation (BA) with bacterial consortium increased the TPHs degradation in aged soil (over 20% compared to natural attenuation - NA). However, co-application of BA with biochar and rhamnolipid higher did not show a statistically prominent synergistic effect. While biochar application facilitated the maintenance of hydrocarbon degrading bacterial consortium in soil, the present study did not identify a direct influence in TPHs degradation. The biochar application in contaminated soil contributed to TPHs adsorption. Rhamnolipid alone slightly increased the TPHs biodegradation with NA, while the combined bioaugmentation treatment with rhamnolipid and biochar increased the degradation between 27.5 and 29.8%. These findings encourage further exploration of combining bioaugmentation with amendment, like biochar and rhamnolipid, for remediating diverse environmental matrices contaminated with complex and aged hydrocarbons.

BacA: a possible regulator that contributes to the biofilm formation of Pseudomonas aeruginosa.

Previously, we pointed out in P. aeruginosa PAO1 biofilm cells the accumulation of a hypothetical protein named PA3731 and showed that the deletion of the corresponding gene impacted its biofilm formation capacity. PA3731 belongs to a cluster of 4 genes (pa3732 to pa3729) that we named bac for "Biofilm Associated Cluster." The present study focuses on the PA14_16140 protein, i.e., the PA3732 (BacA) homolog in the PA14 strain. The role of BacA in rhamnolipid secretion, biofilm formation and virulence, was confirmed by phenotypic experiments with a bacA mutant. Additional investigations allow to advance that the bac system involves in fact 6 genes organized in operon, i.e., bacA to bacF. At a molecular level, quantitative proteomic studies revealed an accumulation of the BAC cognate partners by the bacA sessile mutant, suggesting a negative control of BacA toward the bac operon. Finally, a first crystallographic structure of BacA was obtained revealing a structure homologous to chaperones or/and regulatory proteins.

High mono-rhamnolipids production by a novel isolate Pseudomonas aeruginosa LP20 from oily sludge: characterization, optimization, and potential application.

This study aims to isolate microbial strains for producing mono-rhamnolipids with high proportion. Oily sludge is rich in petroleum and contains diverse biosurfactant-producing strains. A biosurfactant-producing strain LP20 was isolated from oily sludge, identified as Pseudomonas aeruginosa based on phylogenetic analysis of 16S rRNA. High-performance liquid chromatography-mass spectrometry results indicated that biosurfactants produced from LP20 were rhamnolipids, mainly containing Rha-C8-C10, Rha-C10-C10, Rha-Rha-C8-C10, Rha-Rha-C10-C10, Rha-C10-C12:1, and Rha-C10-C12. Interestingly, more mono-rhamnolipids were produced by strain LP20 with a relative abundance of 64.5%. Pseudomonas aeruginosa LP20 optimally produced rhamnolipids at a pH of 7.0 and a salinity of 0.1% using glycerol and nitrate. The culture medium for rhamnolipids by strain LP20 was optimized by response surface methodology. LP20 produced rhamnolipids up to 6.9 g L-1, increased by 116%. Rhamnolipids produced from LP20 decreased the water surface tension to 28.1 mN m-1 with a critical micelle concentration of 60 mg L-1. The produced rhamnolipids emulsified many hydrocarbons with EI24 values higher than 56% and showed antimicrobial activity against Staphylococcus aureus and Cladosporium sp. with inhibition rates 48.5% and 17.9%, respectively. Pseudomonas aeruginosa LP20 produced more proportion of mono-rhamnolipids, and the LP20 rhamnolipids exhibited favorable activities and promising potential in microbial-enhanced oil recovery, bioremediation, and agricultural biocontrol.

In vitro lung epithelial cell model reveals novel roles for Pseudomonas aeruginosa siderophores.

The multidrug-resistant pathogen Pseudomonas aeruginosa is a common nosocomial respiratory pathogen that continues to threaten the lives of patients with mechanical ventilation in intensive care units and those with underlying comorbidities such as cystic fibrosis or chronic obstructive pulmonary disease. For over 20 years, studies have repeatedly demonstrated that the major siderophore pyoverdine is an important virulence factor for P. aeruginosa in invertebrate and mammalian hosts in vivo. Despite its physiological significance, an in vitro, mammalian cell culture model that can be used to characterize the impact and molecular mechanisms of pyoverdine-mediated virulence has only been developed very recently. In this study, we adapt a previously-established, murine macrophage-based model to use human bronchial epithelial (16HBE) cells. We demonstrate that conditioned medium from P. aeruginosa induced rapid 16HBE cell death through the pyoverdine-dependent secretion of cytotoxic rhamnolipids. Genetic or chemical disruption of pyoverdine biosynthesis decreased rhamnolipid production and mitigated cell death. Consistent with these observations, chemical depletion of lipids or genetic disruption of rhamnolipid biosynthesis abrogated the toxicity of the conditioned medium. Furthermore, we also examine the effects of exposure to purified pyoverdine on 16HBE cells. While pyoverdine accumulated within cells, it was largely sequestered within early endosomes, resulting in minimal cytotoxicity. More membrane-permeable iron chelators, such as the siderophore pyochelin, decreased epithelial cell viability and upregulated several pro-inflammatory genes. However, pyoverdine potentiated these iron chelators in activating pro-inflammatory pathways. Altogether, these findings suggest that the siderophores pyoverdine and pyochelin play distinct roles in virulence during acute P. aeruginosa lung infection. IMPORTANCE: Multidrug-resistant Pseudomonas aeruginosa is a versatile bacterium that frequently causes lung infections. This pathogen is life-threatening to mechanically-ventilated patients in intensive care units and is a debilitating burden for individuals with cystic fibrosis. However, the role of P. aeruginosa virulence factors and their regulation during infection are not fully understood. Previous murine lung infection studies have demonstrated that the production of siderophores (e.g., pyoverdine and pyochelin) is necessary for full P. aeruginosa virulence. In this report, we provide further mechanistic insight into this phenomenon. We characterize distinct and novel ways these siderophores contribute to virulence using an in vitro human lung epithelial cell culture model.

Cytotoxic rhamnolipid micelles drive acute virulence in Pseudomonas aeruginosa.

Pseudomonas aeruginosa is an opportunistic human pathogen that has developed multi- or even pan-drug resistance toward most frontline and last resort antibiotics, leading to increasing frequency of infections and deaths among hospitalized patients, especially those with compromised immune systems. Further complicating treatment, P. aeruginosa produces numerous virulence factors that contribute to host tissue damage and immune evasion, promoting bacterial colonization and pathogenesis. In this study, we demonstrate the importance of rhamnolipid production in host-pathogen interactions. Secreted rhamnolipids form micelles that exhibited highly acute toxicity toward murine macrophages, rupturing the plasma membrane and causing organellar membrane damage within minutes of exposure. While rhamnolipid micelles (RMs) were particularly toxic to macrophages, they also caused membrane damage in human lung epithelial cells, red blood cells, Gram-positive bacteria, and even noncellular models like giant plasma membrane vesicles. Most importantly, rhamnolipid production strongly correlated with P. aeruginosa virulence against murine macrophages in various panels of clinical isolates. Altogether, our findings suggest that rhamnolipid micelles are highly cytotoxic virulence factors that drive acute cellular damage and immune evasion during P. aeruginosa infections.

Rhamnolipids: A biosurfactant for the development of lipid-based nanosystems for food applications.

Biosurfactants (surfactants synthesized by microorganisms) are produced by microorganisms and are suitable for use in different areas. Among biosurfactants, rhamnolipids are the most studied and popular, attracting scientists, and industries' interest. Due to their unique characteristics, the rhamnolipids have been used as synthetic surfactants' alternatives and explored in food applications. Besides the production challenges that need to be tackled to guarantee efficient production and low cost, their properties need to be adjusted to the final application, where the pH instability needs to be considered. Moreover, regulatory approval is needed to start being used in commercial applications. One characteristic of interest is their capacity to form oil-in-water nanosystems. Some of the most explored have been nanoemulsions, solid-lipid nanoparticles and nanostructured lipid carriers. This review presents an overview of the main properties of rhamnolipids, asserts the potential and efficiency of rhamnolipids to replace the synthetic surfactants in the development of nanosystems, and describes the rhamnolipids-based nanosystems used in food applications. It also discusses the main characteristics and methodologies used for their characterization and in the end, some of the main challenges are highlighted.

Biosurfactants-based mixed polycyclic aromatic hydrocarbon degradation: From microbial community structure toward non-targeted metabolomic profile determination.

Biosurfactants-based bioremediation is considered an efficient technology to eliminate environmental pollutants including polycyclic aromatic hydrocarbons (PAHs). However, the precise role of rhamnolipids or lipopeptide-biosurfactants in mixed PAH dissipation, shaping microbial community structure, and influencing metabolomic profile remained unclear. In this study, results showed that the maximum PAH degradation was achieved in lipopeptide-assisted treatment (SPS), where the pyrene and phenanthrene were substantially degraded up to 74.28 % and 63.05 % respectively, as compared to rhamnolipids (SPR) and un-aided biosurfactants (SP). Furthermore, the high throughput sequencing analysis revealed a significant change in the PAH-degrading microbial community, with Proteobacteria being the predominant phylum (>98 %) followed by Bacteroidota and Firmicutes in all the treatments. Moreover, Pseudomonas and Pannonibacter were found as highly potent bacterial genera for mixed PAH degradation in SPR, SPS, and SP treatments, nevertheless, the abundance of the genus Pseudomonas was significantly enhanced (>97 %) in SPR treatment groups. On the other hand, the non-targeted metabolomic profile through UHPLC-MS/MS exhibited a remarkable change in the metabolites of amino acids, carbohydrates, and lipid metabolisms by the input of rhamnolipids or lipopeptide-biosurfactants whereas, the maximum intensities of metabolites (more than two-fold) were observed in SPR treatment. The findings of this study suggested that the aforementioned biosurfactants can play an indispensable role in mixed PAH degradation as well as seek to offer new insights into shifts in PAH-degrading microbial communities and their metabolic function, which can guide the development of more efficient and targeted strategies for complete removal of organic pollutants such as PAH from the contaminated environment.

Biocompatible Rhamnolipid Self-Assemblies with pH-Responsive Antimicrobial Activity.

There is an urgent need for alternative antimicrobial materials due to the growing challenge of bacteria becoming resistant to conventional antibiotics. This study demonstrates the creation of a biocompatible pH-switchable antimicrobial material by combining bacteria-derived rhamnolipids (RL) and food-grade glycerol monooleate (GMO). The integration of RL into dispersed GMO particles, with an inverse-type liquid crystalline cubic structure in the core, leads to colloidally stable supramolecular materials. The composition and pH-triggered structural transformations are studied with small-angle X-ray scattering, cryogenic transmission electron microscopy, and dynamic light scattering. The composition-structure-activity relationship is analyzed and optimized to target bacteria at acidic pH values of acute wounds. The new RL/GMO dispersions reduce Staphylococcus aureus (S. aureus) populations by 7-log after 24 h of treatment with 64 µg mL-1 of RL and prevent biofilm formation at pH = 5.0, but have no activity at pH = 7.0. Additionally, the system is active against methicillin-resistant S. aureus (MRSA) with minimum inhibitory concentration of 128 µg mL-1 at pH 5.0. No activity is found against several Gram-negative bacteria at pH 5.0 and 7.0. The results provide a fundamental understanding of lipid self-assembly and the design of lipid-based biomaterials, which can further guide the development of alternative bio-based solutions to combat bacteria.

Structural characterization of a nonionic rhamnolipid from Burkholderia lata.

We present the isolation and structural characterization of a novel nonionic dirhamnolipid methyl ester produced by the bacterium Burkholderia lata. The structure and the absolute configuration of the isolated dirhamnolipid bearing a symmetrical C14-C14 methyl ester chain were thoroughly investigated through chemical degradation and spectroscopic methods including 1D and 2D NMR analysis, HR-ESI-TOF-MS, chiral GC-MS, and polarimetry. Our work represents the first mention in the literature of a rhamnolipid methyl ester from Burkholderia species.