Many notable human pathogens are facultative anaerobes. These pathogens exhibit redundant metabolic pathways and a whole array of regulatory systems to adapt to changing oxygen levels. However, our ...knowledge of facultative anaerobic pathogens is mostly based on fully aerobic or anaerobic cultures, which does not reflect real infection conditions, while the microaerobic range remains understudied. Here, we examine the behavior of pathogenic and nonpathogenic strains of two facultative anaerobes, Escherichia coli and Pseudomonas aeruginosa, during the aerobic‐anaerobic transition. To do so, we introduce a new technique named AnaeroTrans, in which we allow self‐consumption of oxygen by steady‐state cultures and monitor the system by measuring the gas‐phase oxygen concentration. We explore the different behavior of the studied species toward oxygen and examine how this behavior is associated with the targeted infection sites. As a model, we characterize the adaptation profile of the ribonucleotide reductase network, a complex oxygen‐dependent enzymatic system responsible for the generation of the deoxyribonucleotides. We also explore the actions of the most important anaerobic regulators and how these regulators influence bacterial fitness. Our results allow us to classify the different elements that compose the aerobic‐anaerobic transition into reproducible stages, thus showing the different adaptation mechanisms of the studied species.
Aggregates of Pseudomonas aeruginosa form a protective barrier against antibiotics and the immune system. These barriers, known as biofilms, are associated with several infectious diseases. One of ...the main components of these biofilms is alginate, a homo- and hetero-polysaccharide that consists of β-D-mannuronate (M) and α-L-guluronate (G) units. Alginate lyases degrade this sugar and have been proposed as biotherapeutic agents to dissolve P. aeruginosa biofilms. However, there are contradictory reports in the literature regarding the efficacy of alginate lyases against biofilms and their synergistic effect with antibiotics. We found that most positive reports used a commercial crude extract from Flavobacterium multivorum as the alginate lyase source. By using anion exchange chromatography coupled to nano LC MS/MS, we identified two distinct enzymes in this extract, one has both polyM and polyG (polyM/G) degradation activities and it is similar in sequence to a broad-spectrum alginate lyase from Flavobacterium sp. S20 (Alg2A). The other enzyme has only polyG activity and it is similar in sequence to AlyA1 from Zobellia galactanivorans. By characterizing both of these enzymes together with three recombinant alginate lyases (a polyM, a polyG and a polyM/G), we showed that only enzymes with polyM/G activity such as Alg2A and A1-II' (alginate lyase from Sphingomonas sp.) are effective in dissolving biofilms. Furthermore, both activities are required to have a synergistic effect with antibiotics.
Six morpholine-(iso)thiosemicarbazone hybrids HL 1 –HL 6 and their Cu(II) complexes with good-to-moderate solubility and stability in water were synthesized and characterized. Cu(II) complexes ...Cu(L 1–6 )Cl (1–6) formed weak dimeric associates in the solid state, which did not remain intact in solution as evidenced by ESI-MS. The lead proligands and Cu(II) complexes displayed higher antiproliferative activity in cancer cells than triapine. In addition, complexes 2–5 were found to specifically inhibit the growth of Gram-positive bacteria Staphylococcus aureus with MIC50 values at 2–5 μg/mL. Insights into the processes controlling intracellular accumulation and mechanism of action were investigated for 2 and 5, including the role of ribonucleotide reductase (RNR) inhibition, endoplasmic reticulum stress induction, and regulation of other cancer signaling pathways. Their ability to moderately inhibit R2 RNR protein in the presence of dithiothreitol is likely related to Fe chelating properties of the proligands liberated upon reduction.
The coexistence between species that occurs in some infections remains hard to achieve in vitro since bacterial fitness differences eventually lead to a single organism dominating the mixed culture. ...Pseudomonas aeruginosa and Staphylococcus aureus are major pathogens found growing together in biofilms in disease-affected lungs or wounds. Herein, we tested and analyzed different culture media, additives and environmental conditions to support P. aeruginosa and S. aureus coexistence in vitro. We have unraveled the potential of DMEM to support the growth of these two organisms in mature cocultured biofilms (three days old) in an environment that dampens the pH rise. Our conditions use equal initial inoculation ratios of both strains and allow the stable formation of separate S. aureus microcolonies that grow embedded in a P. aeruginosa biofilm, as well as S. aureus biofilm overgrowth when bovine serum albumin is added to the system. Remarkably, we also found that S. aureus survival is strictly dependent on a well-characterized phenomenon of oxygen stratification present in the coculture biofilm. An analysis of differential tolerance to gentamicin and ciprofloxacin treatment, depending on whether P. aeruginosa and S. aureus were growing in mono- or coculture biofilms, was used to validate our in vitro coculture conditions.
Microbial biofilms are complex three‐dimensional structures where sessile microbes are embedded in a polymeric extracellular matrix. Their resistance toward the host immune system as well as to a ...diverse range of antimicrobial treatments poses a serious health and development threat, being in the top 10 global public health threats declared by the World Health Organization. In an effort to combat biofilm‐related microbial infections, several strategies have been developed to independently eliminate biofilms or to complement conventional antibiotic therapies. However, their limitations leave room for other treatment alternatives, where the application of nanotechnology to biofilm eradication has gained significant relevance in recent years. Their small size, penetration efficiency, and the design flexibility that they present makes them a promising alternative for biofilm infection treatment, although they also present set‐backs. This review aims to describe the main possibilities and limitations of nanomedicine against biofilms, while covering the main aspects of biofilm formation and study, and the current therapies for biofilm treatment.
This article is categorized under:
Therapeutic Approaches and Drug Discovery > Nanomedicine for Infectious Disease
Toxicology and Regulatory Issues in Nanomedicine > Toxicology of Nanomaterials
Toxicology and Regulatory Issues in Nanomedicine > Regulatory and Policy Issues in Nanomedicine
Roadmap of challenges and limitations in using nanomedicine against biofilm infections.
Concerns have been raised about the long-term accumulating effects of triclocarban, a polychlorinated diarylurea widely used as an antibacterial soap additive, in the environment and in human beings. ...Indeed, the Food and Drug Administration has recently banned it from personal care products. Herein, we report the synthesis, antibacterial activity and cytotoxicity of novel
,
'-diarylureas as triclocarban analogs, designed by reducing one or more chlorine atoms of the former and/or replacing them by the novel pentafluorosulfanyl group, a new bioisostere of the trifluoromethyl group, with growing importance in drug discovery. Interestingly, some of these pentafluorosulfanyl-bearing ureas exhibited high potency, broad spectrum of antimicrobial activity against Gram-positive bacterial pathogens, and high selectivity index, while displaying a lower spontaneous mutation frequency than triclocarban. Some lines of evidence suggest a bactericidal mode of action for this family of compounds.
, a pathogenic yeast associated with systemic infections, exhibits metabolic adaptability in response to nutrient availability.
We investigated the impact of RPMI glucose supplemented (RPMId), TSB, ...BHI and YPD media on
growth, morphology, susceptibility (caspofungin and amphotericin B), and
virulence (
) in planktonic and biofilm states.
High-glucose media favors growth but hinders metabolic activity and filamentation. Media promoting carbohydrate production reduces biofilm susceptibility. Virulence differences between planktonic cells and biofilm suspensions from the same media shows that biofilm-related factors influence infection outcome depending on nutrient availability. Pseudohyphal growth occurred in biofilms under low oxygen and shear stress, but its presence is not exclusively correlated with virulence.
This study provides valuable insights into the intricate interplay between nutrient availability and
pathogenicity. It emphasizes the importance of considering pathogen behavior in diverse conditions when designing research protocols and therapeutic strategies.
Oleanolic acid (OA) and maslinic acid (MA) are pentacyclic triterpenic compounds that abound in industrial olive oil waste. These compounds have renowned antimicrobial properties and lack ...cytotoxicity in eukaryotic cells as well as resistance mechanisms in bacteria. Despite these advantages, their antimicrobial activity has only been tested in vitro, and derivatives improving this activity have not been reported. In this work, a set of 14 OA and MA C-28 amide derivatives have been synthesized. Two of these derivatives, MA-HDA and OA-HDA, increase the in vitro antimicrobial activity of the parent compounds while reducing their toxicity in most of the Gram-positive bacteria tested, including a methicillin-resistant Staphylococcus aureus-MRSA. MA-HDA also shows an enhanced in vivo efficacy in a Galleria mellonella invertebrate animal model of infection. A preliminary attempt to elucidate their mechanism of action revealed that these compounds are able to penetrate and damage the bacterial cell membrane. More significantly, their capacity to reduce antibiofilm formation in catheters has also been demonstrated in two sets of conditions: a static and a more challenged continuous-flow S. aureus biofilm.
Pseudomonas aeruginosa
is a major pathogenic bacterium in chronic infections and is a model organism for studying biofilms.
P. aeruginosa
is considered an aerobic bacterium, but in the presence of ...nitrate, it also grows in anaerobic conditions. Oxygen diffusion through the biofilm generates metabolic and genetic diversity in
P. aeruginosa
growth, such as in ribonucleotide reductase activity. These essential enzymes are necessary for DNA synthesis and repair. Oxygen availability determines the activity of the three-ribonucleotide reductase (RNR) classes. Class II and III RNRs are active in the absence of oxygen; however, class II RNRs, which are important in
P. aeruginosa
biofilm growth, require a vitamin B
12
cofactor for their enzymatic activity. In this work, we elucidated the conditions in which class II RNRs are active due to vitamin B
12
concentration constraints (biosynthesis or environmental availability). We demonstrated that increased vitamin B
12
levels during aerobic, stationary and biofilm growth activate class II RNR activity. We also established that the
cobN
gene is essentially responsible for B
12
biosynthesis under planktonic and biofilm growth. Our results unravel the mechanisms of dNTP synthesis by
P. aeruginosa
during biofilm growth, which appear to depend on the bacterial strain (laboratory-type or clinical isolate).
The hydrophobic composition of mycobacterial cell walls leads to the formation of clumps when attempting to resuspend mycobacteria in aqueous solutions. Such aggregation may interfere in the ...mycobacteria-host cells interaction and, consequently, influence their antitumor effect. To improve the immunotherapeutic activity of Mycobacterium brumae, we designed different emulsions and demonstrated their efficacy. The best formulation was initially selected based on homogeneity and stability. Both olive oil (OO)- and mineral oil-in-water emulsions better preserved the mycobacteria viability and provided higher disaggregation rates compared to the others. But, among both emulsions, the OO emulsion increased the mycobacteria capacity to induce cytokines' production in bladder tumor cell cultures. The OO-mycobacteria emulsion properties: less hydrophobic, lower pH, more neutralized zeta potential, and increased affinity to fibronectin than non-emulsified mycobacteria, indicated favorable conditions for reaching the bladder epithelium in vivo. Finally, intravesical OO-M. brumae-treated mice showed a significantly higher systemic immune response, together with a trend toward increased tumor-bearing mouse survival rates compared to the rest of the treated mice. The physicochemical characteristics and the induction of a robust immune response in vitro and in vivo highlight the potential of the OO emulsion as a good delivery vehicle for the mycobacterial treatment of bladder cancer.
PDF
