Nanoparticles—Attractive Carriers of Antimicrobial Essential Oils

Microbial pathogens are the most prevalent cause of chronic infections and fatalities around the world. Antimicrobial agents including antibiotics have been frequently utilized in the treatment of infections due to their exceptional outcomes. However, their widespread use has resulted in the emergence of multidrug-resistant strains of bacteria, fungi, viruses, and parasites. Furthermore, due to inherent resistance to antimicrobial drugs and the host defence system, the advent of new infectious diseases, chronic infections, and the occurrence of biofilms pose a tougher challenge to the current treatment line. Essential oils (EOs) and their biologically and structurally diverse constituents provide a distinctive, inexhaustible, and novel source of antibacterial, antiviral, antifungal, and antiparasitic agents. However, due to their volatile nature, chemical susceptibility, and poor solubility, their development as antimicrobials is limited. Nanoparticles composed of biodegradable polymeric and inorganic materials have been studied extensively to overcome these limitations. Nanoparticles are being investigated as nanocarriers for antimicrobial delivery, antimicrobial coatings for food products, implantable devices, and medicinal materials in dressings and packaging materials due to their intrinsic capacity to overcome microbial resistance. Essential oil-loaded nanoparticles may offer the potential benefits of synergism in antimicrobial activity, high loading capacity, increased solubility, decreased volatility, chemical stability, and enhancement of the bioavailability and shelf life of EOs and their constituents. This review focuses on the potentiation of the antimicrobial activity of essential oils and their constituents in nanoparticulate delivery systems for a wide range of applications, such as food preservation, packaging, and alternative treatments for infectious diseases.

Adaptation to an amoeba host leads to Pseudomonas aeruginosa isolates with attenuated virulence

The opportunistic pathogen Pseudomonas aeruginosa , is ubiquitous in the environment, and in humans is capable of causing acute or chronic infections. In the natural environment, predation by bacterivorous protozoa represents a primary threat to bacteria. Here, we determined the impact of long-term exposure of P. aeruginosa to predation pressure. P. aeruginosa persisted when co-incubated with the bacterivorous Acanthamoeba castellanii for extended periods and produced genetic and phenotypic variants. Sequencing of late-stage amoeba-adapted P. aeruginosa isolates demonstrated single nucleotide polymorphisms within genes that encode known virulence factors and this correlated with a reduction in expression of virulence traits. Virulence towards the nematode, Caenorhabditis elegans , was attenuated in late-stage amoeba-adapted P. aeruginosa compared to early-stage amoeba-adapted and non-adapted counterparts. Further, late-stage amoeba-adapted P. aeruginosa showed increased competitive fitness and enhanced survival in amoeba as well as in macrophage and neutrophils. Interestingly, our findings indicate that the selection imposed by amoeba resulted in P. aeruginosa isolates with reduced virulence and enhanced fitness, similar to those recovered from chronic cystic fibrosis infections. Thus, predation by protozoa and long-term colonization of the human host may represent similar environments that select for similar losses of gene function. Importance Pseudomonas aeruginosa is an opportunistic pathogen that causes both acute infections in plants and animals, including humans, and chronic infections in immunocompromised and cystic fibrosis patients. This bacterium is commonly found in soils and water where bacteria are constantly under threat of being consumed by bacterial predators, e.g. protozoa. To escape being killed, bacteria have evolved a suite of mechanisms that protect them from being consumed or digested. Here, we examine the effect of long-term predation on the genotypes and phenotypes expressed by P. aeruginosa . We show that long term co-incubation with protozoa resulted in mutations that resulted in P. aeruginosa becoming less pathogenic. This is particularly interesting as we see similar mutations arise in bacteria associated with chronic infections. Importantly, the genetic and phenotypic traits possessed by late-stage amoeba-adapted P. aeruginosa are similar to what is observed for isolates obtained from chronic cystic fibrosis infections. This notable overlap in adaptation to different host types suggests similar selection pressures amongst host cell types as well as similar adaptation strategies.

Bacterial biofilms predominate in both acute and chronic human lung infections

BackgroundA basic paradigm of human infection is that acute bacterial disease is caused by fast growing planktonic bacteria while chronic infections are caused by slow-growing, aggregated bacteria, a phenomenon known as a biofilm. For lung infections, this paradigm has been thought to be supported by observations of how bacteria proliferate in well-established growth media in the laboratory—the gold standard of microbiology.ObjectiveTo investigate the bacterial architecture in sputum from patients with acute and chronic lung infections.MethodsAdvanced imaging technology was used for quantification and direct comparison of infection types on fresh sputum samples, thereby directly testing the acute versus chronic paradigm.ResultsIn this study, we compared the bacterial lifestyle (planktonic or biofilm), growth rate and inflammatory response of bacteria in freshly collected sputum (n=43) from patient groups presenting with acute or chronic lung infections. We found that both acute and chronic lung infections are dominated by biofilms (aggregates of bacteria within an extracellular matrix), although planktonic cells were observed in both sample types. Bacteria grew faster in sputum from acute infections, but these fast-growing bacteria were enriched in biofilms similar to the architecture thought to be reserved for chronic infections. Cellular inflammation in the lungs was also similar across patient groups, but systemic inflammatory markers were only elevated in acute infections.ConclusionsOur findings indicate that the current paradigm of equating planktonic with acute and biofilm with chronic infection needs to be revisited as the difference lies primarily in metabolic rates, not bacterial architecture.

Microbial Biofilms: Structural Plasticity and Emerging Properties

Microbial biofilms are found everywhere and can be either beneficial or detrimental, as they are involved in crucial ecological processes and in severe chronic infections. The functional properties of biofilms are closely related to their three-dimensional (3D) structure, and the ability of microorganisms to collectively and dynamically shape the community spatial organization in response to stresses in such biological edifices. A large number of works have shown a relationship between the modulation of the spatial organization and ecological interactions in biofilms in response to environmental fluctuations, as well as their emerging properties essential for nutrient cycling and bioremediation processes in natural environments. On the contrary, numerous studies have emphasized the role of structural rearrangements and matrix production in the increased tolerance of bacteria in biofilms toward antimicrobials. In these last few years, the development of innovative approaches, relying on recent technological advances in imaging, computing capacity, and other analytical tools, has led to the production of original data that have improved our understanding of this close relationship. However, it has also highlighted the need to delve deeper into the study of cell behavior in such complex communities during 3D structure development and maturation— from a single-cell to a multicellular scale— to better control or harness positive and negative impacts of biofilms. For this Special Issue, the interplay between biofilm emerging properties and their 3D spatial organization considering different models, from single bacteria to complex environmental communities, and various environments, from natural ecosystems to industrial and medical settings are addressed.

Amphotericin B Polymer Nanoparticles Show Efficacy against Candida Species Biofilms

Purpose: Chronic infections of Candida albicans are characterised by the embedding of budding and entwined filamentous fungal cells into biofilms. The biofilms are refractory to many drugs and Candida biofilms are associated with ocular fungal infections. The objective was to test the activity of nanoparticulate amphotericin B (AmB) against Candida biofilms. Methods: AmB was encapsulated in the Molecular Envelope Technology (MET, N-palmitoyl-N-monomethyl-N,N-dimethyl-N,N,N-trimethyl-6-O-glycolchitosan) nanoparticles and tested against Candida biofilms in vitro. Confocal laser scanning microscopy (CLSM) imaging of MET nanoparticles’ penetration into experimental biofilms was carried out and a MET-AmB eye drop formulation was tested for its stability. Results: MET-AmB formulations demonstrated superior activity towards C. albicans biofilms in vitro with the EC50 being ~30 times lower than AmB alone (EC50 MET-AmB = 1.176 μg mL−1, EC50 AmB alone = 29.09 μg mL−1). A similar superior activity was found for Candida glabrata biofilms, where the EC50 was ~10× lower than AmB alone (EC50 MET-AmB = 0.0253 μg mL−1, EC50 AmB alone = 0.289 μg mL−1). CLSM imaging revealed that MET nanoparticles penetrated through the C. albicans biofilm matrix and bound to fungal cells. The activity of MET-AmB was no different from the activity of AmB alone against C. albicans cells in suspension (MET-AmB MIC90 = 0.125 μg mL−1, AmB alone MIC90 = 0.250 μg mL−1). MET-AmB eye drops were stable at room temperature for at least 28 days. Conclusions: These biofilm activity findings raise the possibility that MET-loaded nanoparticles may be used to tackle Candida biofilm infections, such as refractory ocular fungal infections.

Molecular Mechanisms of Staphylococcus and Pseudomonas Interactions in Cystic Fibrosis

Cystic fibrosis (CF) is an autosomal recessive genetic disorder that is characterized by recurrent and chronic infections of the lung predominantly by the opportunistic pathogens, Gram-positive Staphylococcus aureus and Gram-negative Pseudomonas aeruginosa. While S. aureus is the main colonizing bacteria of the CF lungs during infancy and early childhood, its incidence declines thereafter and infections by P. aeruginosa become more prominent with increasing age. The competitive and cooperative interactions exhibited by these two pathogens influence their survival, antibiotic susceptibility, persistence and, consequently the disease progression. For instance, P. aeruginosa secretes small respiratory inhibitors like hydrogen cyanide, pyocyanin and quinoline N-oxides that block the electron transport pathway and suppress the growth of S. aureus. However, S. aureus survives this respiratory attack by adapting to respiration-defective small colony variant (SCV) phenotype. SCVs cause persistent and recurrent infections and are also resistant to antibiotics, especially aminoglycosides, antifolate antibiotics, and to host antimicrobial peptides such as LL-37, human β-defensin (HBD) 2 and HBD3; and lactoferricin B. The interaction between P. aeruginosa and S. aureus is multifaceted. In mucoid P. aeruginosa strains, siderophores and rhamnolipids are downregulated thus enhancing the survival of S. aureus. Conversely, protein A from S. aureus inhibits P. aeruginosa biofilm formation while protecting both P. aeruginosa and S. aureus from phagocytosis by neutrophils. This review attempts to summarize the current understanding of the molecular mechanisms that drive the competitive and cooperative interactions between S. aureus and P. aeruginosa in the CF lungs that could influence the disease outcome.

The Association of Toxoplasma gondii IgG Antibody and Chronic Kidney Disease Biomarkers

Background: Toxoplasma gondii (T. gondii) is a parasite that infects more than 40 million Americans and causes toxoplasmosis. Most cases of toxoplasmosis are asymptomatic; however, T. gondii is capable of invading organs like the kidney, causing chronic infections and cell destruction. Methods: This study focused on evaluating the association between T. gondii exposure and chronic kidney disease (CKD) using data from the 2009–2010 National Health and Nutrition Examination Survey (NHANES). T. gondii exposure was assessed using Toxoplasma gondii IgG antibody status, and the status of CKD was assessed using the CKD biomarkers. The evaluation of risk rate and population prevalence was performed. In addition, multivariable regression models were used to further investigate this association after adjusting for sociodemographic, anthropometric, behavioral, and clinical covariates commonly associated with kidney dysfunction. Results: The positive T. gondii IgG antibody participants had significantly higher levels of CKD biomarkers, including second albumin-to-creatinine ratio (p = 0.0376), second albuminuria (p = 0.0005), and persistent albuminuria (p < 0.0001) compared to the negative participants. Furthermore, there were statistical associations between T. gondii exposure and the status of CKD (negative vs. positive) (p = 0.0001), and between T. gondii exposure and the CKD stage (negative, stage 1, …, stage 5) (p = 0.0004). Without adjusting for age, the positive T. gondii participants had a significantly higher risk (27% higher) of having CKD than the negative participants (RRcrude = 1.27, 95% CI: 1.09–1.49). The age-adjusted prevalence of CKD was higher among Toxoplasma-positive participants compared to the Toxoplasma-negative participants (10.45 vs. 8.99). T. gondii infection was significantly associated with CKD (OR = 1.40, 95% CI = 1.06–1.84, p = 0.00447) after adjusting for age, gender, race/ethnicity, and BMI. Age was positively associated with CKD (OR = 8.89, 95% CI = 6.31–12.51, p < 0.0001) with the participants 45+ years old being 8.89 times more likely to have CKD than those who are <45 years old, after adjusting for T. gondii infection, gender, race/ethnicity, and BMI. Moreover, positive T. gondii increased the odds of CKD progression (OR = 1.41, 95% CI = 1.07–1.86, p = 0.0424). Conclusions: Positive T. gondii IgG antibody is associated with CKD and the progression of CKD stages. This association is more apparent among older people. Further investigations are needed to examine these findings in different geographical locations and among differentially exposed populations.

Distribution of serotypes and antibiotic resistance of invasive Pseudomonas aeruginosa in a multi-country collection

Background Pseudomonas aeruginosa is an opportunistic pathogen that causes a wide range of acute and chronic infections and is frequently associated with healthcare-associated infections. Because of its ability to rapidly acquire resistance to antibiotics, P. aeruginosa infections are difficult to treat. Alternative strategies, such as a vaccine, are needed to prevent infections. We collected a total of 413 P. aeruginosa isolates from the blood and cerebrospinal fluid of patients from 10 countries located on 4 continents during 2005–2017 and characterized these isolates to inform vaccine development efforts. We determined the diversity and distribution of O antigen and flagellin types and antibiotic susceptibility of the invasive P. aeruginosa. We used an antibody-based agglutination assay and PCR for O antigen typing and PCR for flagellin typing. We determined antibiotic susceptibility using the Kirby-Bauer disk diffusion method. Results Of the 413 isolates, 314 (95%) were typed by an antibody-based agglutination assay or PCR (n = 99). Among the 20 serotypes of P. aeruginosa, the most common serotypes were O1, O2, O3, O4, O5, O6, O8, O9, O10 and O11; a vaccine that targets these 10 serotypes would confer protection against more than 80% of invasive P. aeruginosa infections. The most common flagellin type among 386 isolates was FlaB (41%). Resistance to aztreonam (56%) was most common, followed by levofloxacin (42%). We also found that 22% of strains were non-susceptible to meropenem and piperacillin-tazobactam. Ninety-nine (27%) of our collected isolates were resistant to multiple antibiotics. Isolates with FlaA2 flagellin were more commonly multidrug resistant (p = 0.04). Conclusions Vaccines targeting common O antigens and two flagellin antigens, FlaB and FlaA2, would offer an excellent strategy to prevent P. aeruginosa invasive infections.

Formation, Development, and Cross-Species Interactions in Biofilms

Biofilms, which are essential vectors of bacterial survival, protect microbes from antibiotics and host immune attack and are one of the leading causes that maintain drug-resistant chronic infections. In nature, compared with monomicrobial biofilms, polymicrobial biofilms composed of multispecies bacteria predominate, which means that it is significant to explore the interactions between microorganisms from different kingdoms, species, and strains. Cross-microbial interactions exist during biofilm development, either synergistically or antagonistically. Although research into cross-species biofilms remains at an early stage, in this review, the important mechanisms that are involved in biofilm formation are delineated. Then, recent studies that investigated cross-species cooperation or synergy, competition or antagonism in biofilms, and various components that mediate those interactions will be elaborated. To determine approaches that minimize the harmful effects of biofilms, it is important to understand the interactions between microbial species. The knowledge gained from these investigations has the potential to guide studies into microbial sociality in natural settings and to help in the design of new medicines and therapies to treat bacterial infections.

The Role of Integration Host Factor in Escherichia coli Persister Formation

Persisters are phenotypic variants that survive exposure to antibiotics through temporary dormancy. Mutants with increased levels of persisters have been identified in clinical isolates, and evidence suggests these cells contribute to chronic infections and antibiotic treatment failure.